CHAPTER 20 METHODS OF BREEDING AND THEIR CHARACTERISTICS

In addition to choosing among several objectives has led geneticists to devote con- when tree breeding projects are established, deci- siderable time to establishing arboreta and carrying sions have to be made in regard to specific traits to out species crossability studies. be improved and, what is equally important, the Introduction of forest tree species in the southern method or methods of breeding to be used. Im- United States has long interested landowners and provement in crop plants may be achieved in many . Zon and Brisco (1911) described the suc- different ways, but all methods of breeding do not cess and failures of planted eucalypts in Florida apply equally well to southern pine problems, and dating from about 1878. Gemmer (1931) outlined the advantages of each method have to be balanced plans of the USDA Forest Service for testing intro- against the disadvantages. Also, the tree duced pines in Florida. Locations at which exotic may have an opportunity to use several methods species are tested have been listed in a survey of simultaneously or to combine several methods. genetics and tree breeding research (Dormán 1966). Methods of tree breeding must be evaluated on the Many pine species have been test planted in the basis of genetic and tree breeding principles and, South and some survive, but overall performance most importantly, on the basis of the species and has generally been unsatisfactory. Results of plant- traits with which the work is to be done; cultural ings in Florida were summarized (Kraus 1963), as practices and forest products must also be con- were those in North Carolina (Barber 1953), Mis- sidered. A good method of breeding, in general, sissippi (Schmitt and Namkoong 1965), Louisiana may not be effective for a specific problem. (Grigsby 1969), Georgia (Jones 1966; Reines and Most tree breeding projects have limited time Greene 1956-57), and Texas (Zobel, Campbell, and funds; thus, efficiency or simplicity of various Cech, and Goddard 1956; Long 1973). breeding methods becomes very important, as All these pubhcations describe survival and other pointed out in the chapter on research needs. The information for imported pines in test plots in the chosen method must not only work, it must work southern United States. In no case did survival, better than any alternative method. Execution of a form, height growth, or resistance to insects and breeding plan requiring four generations of an an- diseases equal the general performance of native nual plant is far simpler than four generations of United States pines. Pines from the western forest . Thus, the tree breeder will concen- United States such as lodgepole, ponderosa, and trate on achieving the greatest gain in each genera- Monterey have been tried, as have pines from tion, rather than on only the results after a series of Mexico, British Honduras, Spain, Italy, China, generations. Korea, and Japan. Expectations for many years The individual tree breeding methods discussed were high, but results in this country have been here fit into the organization plan for the South virtually nil. described in chapter 16. Advantages and disadvan- tages of each method are discussed so that they can be used effectively at the proper time and place by the keyman in all this process, namely, the crea- Exotics for Large-Scale Planting in the South tive, applied tree breeder. Also, the more impor- The successful uses of exotics in forest planting in tant technical subjects that should be used to en- South America, South Africa, Australia, New Zea- hance various breeding methods are given. land, and some places in Europe are often cited as examples of the value of this work. It has been pointed out by Thulin (1957) that in Australia insuf- IMPORTATION OF NEW SPECIES ficient high-quaUty trees of Monterey pine occur per acre to justify establishment of seed production Introduction of new species of plants from dif- areas. This may come as a shock to many foresters ferent regions or different countries has been an who like to point to this work as a prime example of important part of breeding programs for many what can be obtained by tree introduction. It is species, and the same can be said of animal - quite true that Monterey pine grows very fast in ing. Nonlocal species have been used in two ways: Australia, but a selective is still to be placed directly into commercial production, or necessary to develop varieties of good quality. to be utilized in hybridization for the purpose of Form of southern pines planted in Australia was so combining one or more desirable traits with local bad that foresters started a tree selection program species. The success achieved by plant and animal in 1938 as part of a seed procurement project to

358 provide better planting stock. While introduction of such as poor form could be selected against in the Fi softwoods has been successful in Australia, there and subsequent generations so that a few undesira- have been no reports of the use of hardwoods to ble traits might not cause a species to be rejected replace eucalyptus. From the standpoint of forest for introduction, especially if it had a very impor- tree breeding in the southern United States, it is tant desirable trait such as resistance to a serious important to realize that, in general, pines exported pest. to other countries on a large scale have been either Complete testing of introduced species requires southern pines or pines from southern latitudes. that they be planted on a variety of sites in each Monterey pine is native in southern California, and climatic zone in the South. Many years must pass Mexican weeping pine is native to Mexico. Thus, before we can be certain what timber yield and the South already has or has test planted the quality will be and whether new species can species that are commonly imported by other coun- withstand exposure to all types of weather condi- tries. Introduction of the two nonsoutheastern tions as well as all types of disease and insect pests. pines, Monterey and Mexican weeping pine, have However, this process could be speeded up by using been unsuccessful in the South because of their inoculation techniques or growth chambers to simu- susceptibility to insect pests and cold damage. late climate conditions at various locations. Therefore, the introduction of species that are superior to the native southern pines does not, at this time, appear at all promising. Introduction of Advantages of Tree Introduction in the South hardwoods might be possible, but the South is well 1. In principle, it may be quicker and cheaper to supplied with these also. Arboreta planting is con- introduce a new species or variety if it exists than tinuing. Certain young eucalyptus trees grow fast to create one through a program in south Florida. Something useful may show up in with local species. the future from this work, particularly as hybridi- 2. Even though not suitable for large-scale field zation material for disease and insect resist- planting, certain species may have genes or blocks ance or adaptability to poor sites. It is for this of genes that can be transferred to a native species reason that performance in arboreta should be care- that is weak in some character such as susceptibil- fully recorded so that records are obtained on form, ity to disease or insect attack. wood quality, and susceptibility or resistance to 3. Introduced species may provide much wider specific pests. This has not been done in the past; latitude in the choice of paternal stock in hybridiza- therefore, no records are available to show why tion than if only local species are used. For exam- certain species failed or if they had certain valuable ple, vigor may occur in crosses between traits. distantly related species, races, or varieties of the There are many ecological factors important in same species, provided, of course, that they will successful plant introduction work, and ignorance cross. or disregard of these factors has been the cause of 4. It is advantageous in hybridization work to most of the failures. Nearly always, trees do best have parental stock readily available. Although pol- when planted on sites and in climates very similar len of some trees such as the pines can be shipped to those of their origin. Seed should be obtained long distances, it is difficult to find competent from the better individuals and races of each people to correctly identify the species, collect the species. In the past, tree introduction has been pollen, and ship it to the tree breeder. most successful in Australia, New Zealand, South Africa, and parts of Europe where native species were few in number or species with wood of the Disadvantages of Tree Introduction in the desirable type—softwoods, for example—did not South exist. Quite the reverse is true of the southern ; in fact, we are blessed by nature with the 1. It is costly and time consuming to test all the best of the pines and have Uttle chance of getting prospective tree species on a variety of sites in the better from somewhere else. South for an adequate period of time to insure reli- able performance data. Introduction for Breeding Stock 2. There is only a slight chance that species growing faster and having higher value wood than Use of non-native tree species to hybridize with local species will be found because there are a great local southern species may be of benefit in some number of southern tree species and many of them tree breeding problems. This is the method used in grow extremely fast. Also, they produce a wide va- breeding bUght-resistant chestnuts, where species riety of high-quality products. There are no large had to be chosen for disease resistance rather than areas in which native species cannot be planted, suitability for forest planting. Undesirable traits although there are few species to choose from near

359 the north and west edges of the southern pine re- gion. 3. Racial variation occurs in most species, par- ticularly those with wide geographic or elevational 3. Foreign species may not have inherent im- range. Environmental pressure is most effective in munity or resistance to local diseases and insect creating ecotypes where variation within the pests. species is large. 4. Foreign species that have a wide geographic 4. Knowledge of racial variation will lead to bet- range may be composed of many strains or races ter understanding of the inherent nature of the which would be very time consuming and costly to species. test plant under a variety of conditions in this coun- try. 5. A racial introduction is genetically variable and would not have some of the undesirable fea- tures of more genetically uniform strains if uni- RACIAL SELECTION AS A METHOD OF formity proves to be undesirable. Strains could be TREE BREEDING fairly uniform in traits, such as form, but geneti- cally variable in resistance to pests. Racial selection does not always insure use of a 6. Racial selection is needed to insure that full strain superior to local strains, because none may advantage is taken of the opportunity to use the exist. In cases like this, selection of the local race is best strains occurring naturally. Thus, racial plus best because it preserves good genetic quality. It is single-tree selection and hybridization within well established in forest genetics literature that a species might be productive of significantly better substantial amount of variation in rate of growth tree types. A knowledge of racial variation is essen- and other traits is common in tree species, espe- tial to proper selection of stock in species hybridiza- cially those with wide geographic ranges. However, tion. in the South, except for species like loblolly and slash pines, we cannot go much beyond this point. Disadvantages of Racial Selection in Tree We know variation exists, but we do not know Improvement exactly how much for each major and minor species or what the environmental factors are that are cor- 1. Races may not exist that are better than local related with races. Neither do we know whether stock, particularly if the area is located within the the difference in traits among races in all species is optimum range of the species. Local selection would continuous or discontinuous. Furthermore, we do preserve the best strain, but it would not give local not know which are the better races of every tree planters any improvement. The benefit of ra- species to plant or use as breeding stock in different cial selection may not be the same over all parts of areas. the range of a species. As discussed in the chapters on geographic and 2. Testing an introduced race is expensive, be- racial variation, racial variation studies have been cause much the same procedure is required that is designed in accordance with different objectives, needed for an introduced species. Also, if the such as to determine: (1) the principles of variation species concerned is widespread geographically, in a species, (2) the better sources of seed to plant in many sources must be tested, each on several sites. specific geographic locations within the range, and Fairly long periods of time are required for good (3) the better sources of seed to plant outside the test of resistance to climatic factors and pests. natural range. Thus, the advantages and limitations 3. Test designs are complicated because of the of racial selection as a method of breeding should be large number of seedlings required for a good sam- evaluated on the basis of the problem to be solved. ple of a racial strain, as compared with those of a plus tree or a hybrid, particularly if intraspecific variation is large. Tree breeders have not been able Advantages of Racial Selection in Tree Im- to establish many racial variation studies where all provement sources of variation (tree-to-tree, stand, and race) are under control. 1. Permits use of strains that may be more suita- 4. Racial variation may not exceed tree-to-tree ble for certain purposes than local stock. If racial variation, especially if the species is inherently var- selection is needed to obtain seed for planting out- iable and geographic range is small. Racial varia- side the natural range, then it is good business to tion in vigor, for example, may be much less in slash determine the best areas from which to obtain it. pine than in shortleaf or loblolly pine. 2. If superior races are found, they can be used 5. Tree-to-tree variation may be large within a promptly in forest planting because a fairly large race; thus seed collection without consideration of volume of seed would be available. Vegetative maternal parent may produce seedlings with a mix- propagation to build up seed orchards for volume ture of genotypes ranging from desirable to unde- production would not be needed. sirable. Uniformity of good phenotypes would be

360 lacking, particularly if selection is desired for sev- of fruit and ornamental plants are of this origin eral traits other than vigor, such as oleoresin yield, (Miller 1954). Other selections are crossed directly wood specific gravity, form, tracheid length, and or used in multiple crosses to obtain further refine- chemical properties. Certain traits may be nega- ment in combinations of desirable traits. Lerner tively correlated, or they may be positively corre- (1958) discussed many aspects of selection in his lated. The difficult problem lies with those that are book on this subject. Hayes et al. (1955) and Allard negatively correlated, such as rapid growth and (1960) have given selection extensive treatment in susceptibility to cold injury. For example, loblolly their books on . pine of southern origin grew rapidly in Ohio but was highly susceptible to cold injury (Woerheide 1959). Breeding Cross-Pollinating Species Shortleaf pine planted in Pennsylvania gave similar results (Aughanbaugh 1950). Mode of reproduction of plants is important in evaluation of breeding methods because it controls STAND SELECTION IN TREE BREEDING the manner in which plants can be crossed, and also, it influences variability among individual Stands of forest trees may form a deme or a plants. In general, self-pollinating plants are genetically different unit within species or races, as homozygous, whereas cross-pollinating plants are described in the chapters on stand and racial varia- heterozygous. However, as stated by Hayes et al, tion. Geographic location within species or race may (1955), differences are not clear and sharp, and be an important factor in creating differences plants can be divided into major subdivisions on a among stands. In Texas, progeny of maternal par- basis of mode of reproduction, such as (1) naturally ents in stands on ridges and upper slopes were self-pollinated, (2) often cross-pollinated, (3) natur- more drought resistant than offspring of trees in ally cross-pollinated, and (4) dioecious (male and stands on lower slopes. Evidently, moisture supply female plants). is critical in this area, and there is strong selective The southern pines, as shown in the chapter on pressure for drought resistance. In contrast, slash sexual reproduction, are naturally cross-pollinating pine seedlings from trees on wet and dry sites did species, although a small amount of natural selfing not differ in a Florida study, where summer rainfall occurs in stands. Controlled selfing is possible, but is high. self-compatibility and depression vary It is difficult at this time to set up detailed re- widely among individual trees. Thus, methods for quirements for stands to be converted to seed pro- breeding cross-pollinating species are most impor- duction areas, but the same general procedures can tant for pines, but those for self-pollinating species be followed that are used when selecting plus trees. should not be overlooked. Individual trees may be Stands with an unthrifty appearance, trees of gen- strongly male or female, and dioecism may be a erally poor form, and with a fairly high incidence of factor in breeding, particularly in selection of indi- pest attacks are poor candidates for seed produc- vidual trees. tion areas. To ignore signs of questionable inherent Ways of breeding cross-pollinating plants are: (1) quality might very well defeat the purpose of the mass selection, (2) selection of seed orchard clones, project, which is to designate a stand of as high (3) recurrent selection, (4) synthetic varieties, (5) genetic quality as possible, a stand to be managed , and (6) nursery selection. Each for seed production. method is briefly described and the more important Inasmuch as little is known about stand selection technical aspects given. as a discrete method of breeding, no attempt will be made to give advantages and disadvantages listed for other methods. Mass Selection This method involves making a choice among trees for purposes of seed collections. Only the SINGLE-TREE SELECTION IN TREE maternal parent of the seed is known. Trees are BREEDING chosen for high vigor plus good stem form, good crown form, and freedom from attack by pests. Selection is defined as choosing certain individu- Collection of seed from trees in seed production als with desirable traits from a population. It is the areas, natural stands or , and desirable system by which use is made of the best types geographic locations is a form of mass selection. produced naturally, either for commercial produc- Mass selection is an improvement over no control tion or for further selective breeding. Sometimes, of the maternal parent. Seed of poor maternal and natural variants differ sufficiently from average racial phenotypes is avoided, but gains in wood trees to be placed in production as clones without yield and quality are lower than those from more further modification. Many horticultural varieties intensive breeding methods.Reliable estimates of

361 gain are difficult to obtain because of the difficulty effective method of breeding for southern pines. of choosing adequate control stock, numerous en- Highly important gains in wood yield per acre plus vironmental factors that influence yield and vary improvement in individual traits, such as stem and from year to year, and rigor of selecting trees; also, crown form, natural , wood specific gravity, the large number and combinations of traits to be and resistance to pests, have been obtained. Large evaluated cause difficulty. Wide variability among expenditures for clonal seed orchards in the South- trees is maintained from seed obtained ern States were justified on the basis of evidence by mass selection, but there should be fewer trees from crossing carefully selected plus trees of vari- of poor form and high susceptibihty to certain ous species. pests. Mass selection should be used when small Additional improvements in performance seem amounts of seed are needed or until seed orchards possible because the upper limits to selection and come into production. It can, moreover, be used in hybridization have not been reached. Much re- countries unable to conduct tree improvement proj- search in creative breeding techniques is needed in ects. Mass selection insures use of the best seed this subject to guide practice, which is becoming known, but no controlled crossing or creative highly intensive. breeding is involved. In intraspecific hybrids, family means are im- Application of mass-selection techniques in each proved, but for certain economically important tree species requires a knowledge of subjects such traits, wide differences remain among full sibs. as phenotypic variation, geographic variation, tree Since the genetic base narrows with each genera- selection, racial selection, and random mating. tion, a very broad base is needed at the start if the process is to continue. Just how broad we cannot Selection of Seed Orchard Clones say until the effects of inbreeding in relation to selection and crossing are known. Also, the number Selection of stock for seed orchard establishment of clones required for a seed orchard should be is perhaps one of the quickest and most effective estimated before breeding starts. ways of gaining genetic improvement in a few im- Breeding by recurrent selection makes use of portant traits in southern pines on a large scale in a information from subjects such as tree selection, short time. Trees developing from grafted stock racial selection, breeding for multiple traits, ge- cross-pollinate with others and produce offspring netic disassortative mating, and phenotypic disas- with a higher level of genetic uniformity than those sortative mating. obtained through mass selection. The offspring will not be uniform to a high degree as would clonal plantations, but this is quite desirable from the Synthetic Varieties standpoint of maintaining genetic diversity in re- A synthetic variety is made up of genotypes sistance to pests as insurance against epidemics. which have previously been tested for their ability There is little reason to fear that uniformly to produce superior progeny when crossed in all straight-stemmed trees with well-formed crowns combinations. This method differs from other and wood of desirable quality will not be genetically breeding methods that are defined on the basis of diverse in regard to pest resistance or adaptabiUty. how populations are created. Allard (1960, p. 47) While genetic diversity is important, it is not com- writes, "A synthetic variety is one that has been plete protection against pests. One must remember synthesized from all possible intercrosses among a that genetic diversity has not protected native number of selected genotypes; thereby, a popula- American tree species from chestnut blight, white tion is obtained that is propagated subsequently pine blister rust, or disastrous outbreaks of various from open-pollinated seed." In mass selection, the insects. next generation is propagated from a composite of the seed from phenotypically desirable plants Recurrent Selection selected from the source population, according to Allard (1960). The method consists of selecting parents and The term variety is used to indicate trees with crossing them or their selfed progeny to produce special characteristics but may not be named or populations for reselection (Penny et al 1963). Ad- "taxonomic" varieties. Southern pine breeding is ditional or recurrent cycles of selection can be made leading toward a gradual and sustained improve- as long as satisfactory improvement continues. ment in performance rather than production of Stonecypher (1969a) discussed the general use of named varieties—unlike farm crops that are repro- recurrent selection in tree breeding and outlined duced true to name in order to maintain uniformity the steps in phenotypic and genotypic recurrent and meet seed certification requirements. selection (fig. 202). Breeding synthetic varieties utilizes trees and Recurrent selection is the most important and clones produced under other breeding methods

362 BASE BASE POPULATION POPULATION

selection selection

INTERBREED NTERBREED INTERBREEDING PROGENY SELECTS OF S1 SEED TESTED SELECTS SELECTS / 1 SEED SI

PLANTATIONS First Generation RECORDS OF selection PAREN TAGE

INTERBREEDING selection OF SELECTS INTERBREED I NTERBREEC PROGENY S2|SEED SELECTS TESTED SELECTS

PLANTATIONS SEED S2

selection Second Generation INTERBREEDING RECORDS OF OF PA R E N T A G E SELECTS 1 REPEAT CYCLE PHENOTYPIC RECURRENT SELECTION GENOTYPIC RECURRENT SELECTION

Figure 202.—Diagrams of recurrent selection breeding methods. In phenotypic recurrent selection, the selections are recombined and are the base for the next cycle. In genotypic recurrent selection, records of parentage are required, and selections are based on performance of relatives. (Stonecypher 1969a) such as backcrossing and hybridization. Involved in region. It could also transfer pest resistance to breeding synthetic varieties are tree selection, ra- strains that have high vigor and good form. cial selection, phenotypic assortative mating, and Backcrossing requires a knowledge of tree selec- genetic disassortative mating. tion, racial selection, species selection, genetic dis- assortative mating, breeding for multiple traits, Backcrossing and inbreeding. The method consists of recurrent crossing of offspring to the more desirable parent while selec- Nursery Selection tion is practiced for the characters being transfer- This method involves careful inspection of large red from the donor parent. Thus, it may be a form numbers of seedlings while still in the nursery bed of species hybridization but could be used with and also involves outplanting the most vigorous for trees within species if there were no true species to use as breeding stock or for seed orchards. The serve as donor parents. Backcrossing has been little method has several advantages as well as disadvan- used in pine breeding but could become important tages, although actual tests have not developed to as the use of intensive breeding schemes and sys- the point where we can be certain about many of tems develop. The method would produce trees them. with a very good combination of traits, but it would One advantage of nursery selection is that a large at the same time narrow the genetic base. The number of plants can be examined in a short time, method might be useful in transferring traits of as compared with that required to make surveys for vigor to low-vigor trees that possess the asset of superior trees in wild stands or plantations. And high cold resistance, such as pitch and shortleaf the seedlings selected for outstanding vigor may be pines in the northern part of the southern pine inherently superior. If the nurseries are large, sev-

363 eral hundred individual seedlings may be selected lings of plus trees. in a short time. Studies of the relationship between 3. Single-tree selection plus hybridization within seed weight or size and seedling growth show that the species permits increasing one or more traits ?¿ed weight has some effect on seedling size (Brown without introducing undesirable traits that may and Goddard 1959) but that it is quite small com- occur in a separate species. pared with other factors. Thus, seedlings ap- 4. Single-tree selection, or recurrent selection, parently have different inherent growth enables the tree breeder to obtain improvement capabilities that may continue for many years, as where no superior races occur or where there are shown by results of studies in grading nursery no suitable species with which to hybridize. stock. Tests now being made will show what pro- 5. Improvement is possible in traits that are dif- portion of these continue their habit of rapid ficult to evaluate economically—branch diameter, growth (Ellertsen 1955; Zobel et al. 1957). for example—by including the trait along with One disadvantage of nursery selection is the fact others in selection criteria. that environmental factors in the nursery, such as 6. Progeny of single trees, or trees in stands early germination, soil differences, or other condi- established with seed from seed production areas, tions, may be about equally responsible for the should be of fairly good quality. rapid growth of certain seedlings; thus a large error is involved in selecting plants that are truly out- Disadvantages of Single-Tree Selection in standing in vigor. Another disadvantage is that, as yet, we cannot estimate stem or crown form charac- Tree Improvement teristics of mature trees from young seed- 1. Some important traits may not vary within lings. Since undesirable features, such as stem the species or not vary widely enough to permit crook, high branch angle, long branch length, poor selection for improved varieties. For example, re- natural pruning, and probably stem taper, vary in sistance to tip moth in loblolly or shortleaf pines is the species and are under rather strong genetic nonexistent or so low that it does not appear to be control, the method involves an important weak- feasible to select for it. ness. Also, no estimate of wood quality such as 2. Rigid selection may lead to loss of vigor specific gravity, fibril angle, or cellulose content through inbreeding, and losses to pests might in- can be made without sacrificing the seedling. In crease because genetic diversity is reduced. some breeding programs, such considerations 3. Selection for a large number of traits is very would be very important; in others, they would not. difficult because of the survey work required to find The same objection would apply to selection for individuals outstanding in a great many traits. disease resistance. In addition, even though some However, this obstacle applies to interspecific hy- outstanding seedlings may be selected, several bridization, too. years of growth must accumulate before the tree 4. Selection for a combination of certain traits breeder can be sure he has chosen a desirable may be difficult because they are negatively corre- genotype. So, unless selection is highly accurate, lated. several years of testing are required for rating the individual trees, just as are required for estimating the genotype of mature trees. Our inability to accu- SPECIES HYBRIDIZATION AS A rately predict tree growth and areawise wood vol- ume yield is an important factor in progeny testing. METHOD OF TREE BREEDING Hybridization of species has been a powerful tool Advantages of Single-Tree Selection in Tree for the plant breeder. Genera composed of closely Improvement related but varying species have permitted the plant breeder to create an almost unlimited number 1. Since most species vary genetically, especially of combinations of traits in different varieties. southern pines, use of the best stock occurring Thus, collection of species and varieties, although naturally offers an opportunity to obtain an im- they are distributed worldwide, followed by provement over wild stock. The improvement in cataloging their traits, has been an important part some traits is more than in others; the variation in of many plant breeding programs. This technique some species is greater than in others. has been particularly effective in breeding cereal 2. Single-tree selection can easily be used by sil- grains and other important crops. Hybrid vigor, the viculturists, since pollen handling and controlled goal of many plant breeders, is not fully under- pollinations may not be required for simple selec- stood, as indicated by Gowen (1952) in , tion methods such as seed collection from plus and does not appear to occur in southern pine hy- trees, seed production area establishment, and seed brids. orchards establishment with open-pollinated seed- The term hybridization has been used, in gener-

364 al, to denote crossing between plants of different in some cases this may limit the number of combina- species. A very well known exception to this is tions possible. hybrid corn, which is not crossed between species 2. Compatible species and species adapted to a but between different lines. As Duffield and Snyder specific geographic area may not vary widely in a (1958) point out, interspecific hybridization refers large number of economically important traits. to crosses between species, intraspecific hybridiza- Thus, improvement is limited—wood quality tion to crosses between individuals within a species, characteristics among southern pine species, for and interracial hybridization to crosses between example, do not vary widely. members of populations within species. These are very useful terms and are needed to describe fully 3. Although various species may be compatible, the work that has been done in hybridization of some may have undesirable traits; thus, first- or forest trees. second-generation hybrids are not suitable for Hybridization work with poplars (Schreiner planting, and several generations of trees may have 1958), with pine (USDA Forest Service 1948a; to be produced before the desired combination of Righter and Duffield 1951; Wright and Gabriel traits is obtained. This means that improvement 1958; Wright 1959), and with spruce and other would be very slow. Hybridization requires ma- species (Wright 1953) seems to be based largely on nipulation of all the traits, good as well as bad, of random crossing of all different species. This ap- two species, which is more difficult than improving proach provides a pattern of crossability that will traits of a single species. be very useful in future work. Also, it is highly 4. Seed set in some hybrids may be very low. commendable from the statistical point of view be- 5. Seed may be difficult to produce in large quan- cause it is certainly an unbiased sample—all species tities because species may bloom at different times. being crossed. However, yield of improved strains Seed orchards of vegetatively propagated material for wide-scale commercial use has been low. Many from the Fi or Fg generation, however, may be species of some genera are not important commer- feasible. cially, and crosses between them, or between them 6. Species may be only partly compatible, and and more valuable species, cannot be expected to many undesirable individual trees may be produced yield anything very useful. that cannot be identified until after they have been When using hybridization, knowledge of such outplanted several years. Loblolly x slash hybrids subjects as selection of trees, races, or species is in Georgia, for instance, have many aberrant forms. important, as are compatibility, phenotypic assort- ative mating, genetic disassortative mating, inher- 7. Segregation in the F2 or subsequent genera- itance, heritability of traits, and breeding for mul- tions after outplanting may result in the establish- tiple traits. ment in the forest of vigorous trees with undesira- ble traits. 8. Hybrids of species or races that occur in dif- Advantages of Hybridization in Tree ferent geographic areas or climatic areas require a Improvement fairly long period of observation before their resist- ance to climatic factors and pests can be ascer- 1. An extremely large number of genie combina- tained for the specific areas in which they are to be tions is possible because variation among species is grown. Wide variation within species broadens the often greater than among trees or races within opportunities for hybridization, but it complicates species. Furthermore, species not only vary but the testing of hybrids as well. may be composed of many races and individual trees with outstanding traits. This permits a very 9. Insufficient evidence on heritability of traits is wide selection of material. available upon which to base estimates of charac- 2. Hybrid vigor may occur in species crosses. teristics of a hybrid of certain species. Eventually, 3. It may be possible to obtain progeny with we will know more about traits that are dominant, traits that do not occur in either parent. intermediate, or recessive, which will aid greatly in 4. Hybrids of many tree species are fertile. making proper choices of parental stock. 5. It may be possible to cross incompatible 10. Performance of a hybrid may be strongly in- species by using still other species as a bridge, thus fluenced by the individual maternal or paternal transferring genes of blocks of genes. parents that were used; hence, duplication of re- sults may be difficult unless the race and genotype of the parents are given. Disadvantages of Hybridization in Tree 11. Silviculturists who have no special training in Improvement pollination techniques will find hybridization dif- 1. Crossing is limited to compatible species, and ficult.

365 POLYPLOID AND MUTATION reviewed by Brown (1967), but the work has not BREEDING sufficiently advanced so that the advantages and disadvantages can be compared. The method re- The effects of multiple sets of chromosomes quires growth of gametophytic tissue followed by (polyploidy) and heritable changes in genes or chromosome doubling and the formation of com- chromosomes (mutations) on growth and mor- plete plants. As a result, a homozygous line would phological traits were reviewed in an earlier chap- be established that could not otherwise be created ter. Changes such as these may occur naturally, but without many generations of selfing that might be they can also be induced artificially and, con- extremely costly and technically difficult to pro- sequently, can be classed as methods of plant breed- ing. duce. Nei (1963) recommended haploid breeding for forest trees. The suggestion was based on the as- sumptions that heterosis was possible in crosses of inbred lines and that less time was required to Methods of Inducing Mutations produce the inbred lines by the haploid method than Mutations may be induced by several methods others. But these assumptions are question- having different effects (Briggs and Knowles 1967). able. Burk et al (1972) demonstrated that homozy- One group of materials brings about changes as a gous, true-breeding tobacco (Nicotiana tabacum) result of ionizing radiation, the types most com- strains could be created in one generation. They monly used being X-rays, gamma rays, neutrons, cultured anthers of hybrids to produce haploid beta particles, alpha particles, and protons or plants and then doubled the chromosomes of the deuterons. Ionizing radiation can cause gene muta- haploids by colchicine treatment. Progenies of dou- tion; chromosome, chromatid, or subchromatid bled plants of the same clone did not differ in traits. aberrations; changes in chromosome number; inhi- Although the authors claimed that the results show bition of cell division; induction of mitotic activity; the effectiveness of the method for creating death of nuclei or cells; partial or complete sterility; homozygous lines, they tested it only with tobacco. retardation or stimulation in growth rate; and the In tobacco, it is extremely easy not only to grow induction of abnormal growth. An additional type of haploid plants but to double them by colchicine mutagenic agent is ultraviolet radiation; and com- treatment. Thus, practicality of results with to- pared with X-rays, ultraviolet radiation produces bacco may differ sharply from those obtained with more gene mutations relative to chromosome aber- other species. Kimber and Riley (1963) concluded rations. The last group of materials is made of from a review of the subject of haploids that the chemicals such as alkylating agents, urethane, the greatest obstacle to their use was the lack of alkaloids, the peroxides, formaldehyde, and sub- techniques for creating them when needed. Stettler stances related to nucleic acid and nitrous acid. and Howe (1966) described problems encountered It is apparent that mutation and polyploid breed- in producing haploids in Populus, Haploid breeding ing are complex methods because of the large required that a choice be made among wild trees or number of agents, difficulties in using them, differ- individuals in families for trees with which to begin ences in effects, differences in genetics of various the process. Thus, a knowledge of variation among plants, and the large number of parts of the plants trees is essential. to be treated (pollen, seed, twigs, buds). Advantages of Polyploid and Mutation Breeding in Trees 1. Plants with unusual numbers of chromosomes Mutation breeding with forest trees has followed or changed genes or chromosomes may differ the general methods used with other crops but greatly from natural trees in valuable traits. without notable success. As Gustafsson (1960) and 2. Combinations of traits such as resistance to Khoshoo (1959) point out, polyploidy and mutants several pests may have to be induced because no are rather rare in gymnosperms but more frequent resistant stock exists, and, thus, there are no op- in angiosperms. In Russia, it has been pointed out portunities for breeding. that polyploid occur very seldom in nature 3. Highly uniform strains of plants may be pro- and with rare exceptions are of low vitality duced by haploid breeding. (Matskevich 1959). In his review of breeding methods for forest trees, Brewbaker (1967) was not optimistic about the possibilities of induced poly- Disadvantages of Polyploid or Mutation ploids or mutations, and neither was Cech (1963). Breeding The techniques of breeding haploids have been 1. The processes cannot be directed; thus, an

366 Table 9.—Estimated relative effectiveness of tree-breeding methods such as racial selection, single-tree selection, and hybridization in improving various traits in major southern pines

Traits to be improved Method of breeding : Racial selection Singlertree Hybridization selection Oleoresin yield Poor Good Poor Wood specific gravity Poor Good Poor Rate of growth Good Good Good Tracheid length Poor Good Poor Stem straightness Poor Good Poor Natural pruning ability Poor Good Poor Crown form Poor Good Poor Resistance to drought Good Fair Poor Resistance to fusiform rust Good Good Good Resistance to littleleaf disease Good Good Good Resistance to glaze damage Fair Fair Fair extremely large number of plants must be treated number of advantages and disadvantages. At the and grown in test plots. present time, it can be said that selective breeding 2. Costs of evaluating treated plants or tissue within species combined with racial selection is the are very high. most effective method for obtaining improved 3. Most changes are harmful rather than benefi- strains of softwoods and hardwoods. Of the two cial, which reduces the chances of creating im- methods of breeding, selection and crossing within proved trees but improves the chances of creating species will give more improvement than selection deformed trees. of races. This comparison is based on results of 4. Since vegetative propagation in southern studies to date showing that tree-to-tree variation pines is not easy, a few chance individuals would is greater in economically important traits than is not be very useful. racial variation in certain species, and that tree 5. The methods seem very low in productivity introduction and hybridization are not very promis- per unit of time and effort in comparison to alterna- ing at present for general use. Hybridization for tive approaches to breeding, such as selection and specific objectives will be of value in the future. hybridization, because of the presence of a very On a basis of the results from variation and in- large amount of natural variability within and heritance studies to date, the relative effectiveness among the southern pine species. of the three main methods of tree breeding for 6. The absence of useful variants in natural popu- improving some of the various traits in southern lations is evidence that producing them artificially pines is given in table 9. Similar tables could be might be very difficult. prepared for each species and for various types of 7. Techniques for using various methods on a racial selection, single-tree selection, or hybridiza- large scale have not been developed enough so that tion. tree breeders can use them without an investment From the discussion in the sections on variation of time and expense in developing them. and inheritance in southern pines and on methods of 8. Highly uniform strains of forest trees might tree breeding, it is apparent that great diversity in not be silviculturally desirable because of the risk of genetic material occurs in southern tree species and losses to pests, but their value in comparison to that, under certain conditions, various methods of costs should be studied by researchers. breeding can cause dramatic changes. It is appar- ent, also, that methods of breeding cannot be apphed indiscriminately; consequently, the great need now in southern tree breeding is to define DISCUSSION carefully the specific problems and bring to bear on Tree introduction, racial selection, stand selec- them the most specialized and effective method or tion, single-tree selection, and hybridization have a combination of methods to fit the particular case.

367 Tree breeding

Tree breeding is the application of genetic, reproductive biology and economics principles to the genetic improvement and management of forest trees. In contrast to the selective breeding of livestock, arable crops, and horticultural flowers over the last few centuries, the breeding of trees, with the exception of fruit trees, is a relatively recent occurrence. A typical forest tree breeding program starts with selection of superior phenotypes (plus trees) in a natural or planted forest. This application of mass selection improves the mean performance of the forest. Offspring is obtained from selected trees and grown in test plantations that act as genetic trials. Based on such tests the best genotypes among the parents can be selected. Selected trees are typically multiplied by either seeds or and seed orchards are established when the preferred output is improved seed. Alternatively, the best genotypes can be directly propagated by cuttings or in-vitro methods and used directly in clonal plantations. The first system is frequently used in pines and other conifers, while the second is typical in some broadleaves (poplars, eucalypts and others). The objectives of a tree breeding program range from yield improvement and adaptation to particular conditions, to pest- and disease-resistance, wood properties, etc. Currently, tree breeding starting to take advantage of the fast development in plant genetics and genomics.

Optimization Tree breeders make efforts to get their operation efficient by optimising tree breeding. Scientists develop tools aimed at improvement of the efficiency of tree breeding programmes. Optimising can mean adapting strategies and methods to certain species, group of populations, structure of genetic variation and mode of inheritance of the important traits to obtain the highest benefit per unit of time. Optimising is usually carried out at the following levels: breeding strategy (appropriate intensity of breeding, breeding population structure and size, plan for maintenance of genetic diversity), breeding methods (mating type, testing and selection methods, testing population size and time) and deployment methods of the genetically improved material (seed orchards[1] and clonal : genetic contribution, size). Computer simulators are frequently used: stochastic – based on defined and random algorithms; and deterministic – based on defined algorithms. Selection strategies have been compared for annual progress in long-term breeding at a given annual cost considering genetic gain, gene diversity, cost components, and time components. For Norway spruce it seems favourable to clone full sib families and then select based on clonal performance[2] while for Scots pine a two-stage strategy seems best, first phenotypic pre-selection and then progeny-testing the selections.[3]

Tree improvement

A genetically variable population and a method of selecting genetically superior individuals provide the basis for tree improvement by breeding. In essence, a tree improvement program sets out to isolate and evaluate the genetic component of variation in one or more characters of interest. In the simplest procedure, cycles of selection reduce the available population in a particular direction to enhance desirable traits, then breeding from selections to expand the population with improved characteristics. Breeding strategies vary with species and objectives, but all use mating designs to generate information and new material. Choice of a suitable breeding strategy and mating design is a key decision in any breeding program. Kiss (1986)[4] used a 2-level design in British Columbia to study variation within and between separate populations of white spruce, both within British Columbia and from eastern North America. The breeding program for white spruce initiated in 1986 by the Canadian Forestry Service in the Maritimes employed 2 kinds of mating: polycross, to test clones for general combining ability; and pair- mating, to generate material for second generation selections (Fowler et al. 1988).[5] Newton’s (2003)[6] systematic review of yield responses of white spruce and 3 other North American conifers to forest tree improvement practices indicated that correct provenance-progeny selection could yield juvenile height growth gains of about 12% at 20 years for white spruce, and a corresponding merchantable productivity (mean annual merchantable volume increment) gain of 26% at 50 years for plantations established at nominal initial densities on medium-to-good quality sites. Also, preliminary estimates derived from individual case studies indicated that first generational selection strategies for white spruce could increase merchantable productivity by approximately 20% at 45 years. Anthesis is the period during which a flower is fully open and functional. It may also refer to the onset of that period.The onset of anthesis is spectacular in some species. In Banksia species, for example, anthesis involves the extension of thestyle far beyond the upper perianth parts. Anthesis of flowers is sequential within an inflorescence, so when the style and perianth are different colours, the result is a striking colour change that gradually sweeps along the inflorescence.Flowers with diurnal anthesis generally are brightly colored in order to attract diurnal insects, such as butterflies. Flowers withnocturnal anthesis generally are white or less colorful, and as such, they contrast more strongly with the night. These flowers typically attract nocturnal insects including many moth species. Ploidy Breeding

An organism, in which number of complete chromosome sets is more than the diploid number is called polyploid. It is usual practice to express the chromosome number of an organism in diploid, symbolically represented by 2n. In all sexually propagated crops, such expression is of specific significance - because pairs of chromosomes indicate the number of chromosomes in each genome. The organisms are able to maintain chromosome numbers over generations due to the fact that gametes have n number of chromosomes, while all other cells of the organisms have 2n number of chromosomes.

However, some organisms have more than 2n chromosomes. They are called polypliods. Polyploids may be triploids (3n), tetraploids (4n), hexaploids (6n). When polyploidy occurs within a species it is called autopolyploidy and when polyploidy occurs due to chromosomes from two species it is called allopolyploidy. Polyploidy may be caused by  fertilization of an egg by more than one sperm

 failure of meiosis during gamete formation.

When number of chromosome is odd (except 1), the condition is called aneuploidy, while the condition with even numbers (2, 4, 6, 8) is termed as euploidy.

Polyploidy can artificially be introduced by a chemical called colchicine or by crossing diploids with tetraploids.

Although triploids exhibit sterility, polyploidy is useful in some ways due to some advantages. For e.g.,  In some plants like apple, pear the fruit size is bigger in triploids.

 In sugar beet, high sugar content is associated with triploidy.

Ploidy breeding involves hybridization and selection of plants / varieties /species exhibiting various polyploidy. A great deal of success in wheat breeding was achieved, when origin of hexaploid wheat was discovered. Hexaploid wheat was a result of sequence of natural crossing and chromosome doubling. In the study of crop evolution, many such instances are recorded in crops like potato, paddy and sugarcane.

Wheat is the best example of role of polyploidy in crop improvement.

Breeding Methods in Crop Plants

Classification of crop plants based on mode of pollination and mode of reproduction

Mode of pollination and Examples of crop plants reproduction

Self Pollinated Crops Rice, Wheat, Barley, Oats, Chickpea, Pea, Cowpea, Lentil, Green gram, Black gram, Soybean, Common bean, Moth bean, Linseed, Sesame, Khesari, Sunhemp, Chillies, Brinjal, Tomato, Okra, Peanut, Potato, etc.

Corn, Pearlmillet, Rye, Alfalfa, Radish, Cabbage, Cross Pollinated Crops Sunflower, Sugarbeet, Castor, Red clover, White clover, Safflower, Spinach, Onion, Garlic, Turnip, Squash, Muskmelon, Watermelon, Cucumber, Pumpkin, Kenaf, Oilpalm, Carrot, Coconut, Papaya, Sugarcane, Coffee, Cocoa, Tea, Apple, Pears, Peaches, Cherries, grapes, Almond Strawberries, Pine apple, Banana, Cashew, Irish, Cassava, Taro, Rubber, etc. Often Cross Pollinated Crops Sorghum, Cotton, Triticale, Pigeonpea, Tobacco.

BREEDING METHODS IN CROP PLANTS

SELF POLLINATED CROPS

Mass selection

In mass selection, seeds are collected from (usually a few dozen to a few hundred) desirable appearing individuals in a population, and the next generation is sown from the stock of mixed seed. This procedure, sometimes referred to as phenotypic selection, is based on how each individual looks. Mass selection has been used widely to improve old “land” varieties, varieties that have been passed down from one generation of farmers to the next over long periods.

An alternative approach that has no doubt been practiced for thousands of years is simply to eliminate undesirable types by destroying them in the field. The results are similar whether superior plants are saved or inferior plants are eliminated: seeds of the better plants become the planting stock for the next season.

A modern refinement of mass selection is to harvest the best plants separately and to grow and compare their progenies. The poorer progenies are destroyed and the seeds of the remainder are harvested. It should be noted that selection is now based not solely on the appearance of the parent plants but also on the appearance and performance of their progeny. Progeny selection is usually more effective than phenotypic selection when dealing with quantitative characters of low heritability. It should be noted, however, that progeny testing requires an extra generation; hence gain per cycle of selection must be double that of simple phenotypic selection to achieve the same rate of gain per unit time.

Mass selection, with or without progeny test, is perhaps the simplest and least expensive of plant- breeding procedures. It finds wide use in the breeding of certain forage species, which are not important enough economically to justify more detailed attention.

Pure-line selection Pure-line selection generally involves three more or less distinct steps: (1) numerous superior appearing plants are selected from a genetically variable population; (2) progenies of the individual plant selections are grown and evaluated by simple observation, frequently over a period of several years; and (3) when selection can no longer be made on the basis of observation alone, extensive trials are undertaken, involving careful measurements to determine whether the remaining selections are superior in yielding ability and other aspects of performance.

Any progeny superior to an existing variety is then released as a new “pure-line” variety. Much of the success of this method during the early 1900s depended on the existence of genetically variable land varieties that were waiting to be exploited. They provided a rich source of superior pure-line varieties, some of which are still represented among commercial varieties. In recent years the pure-line method as outlined above has decreased in importance in the breeding of major cultivated species; however, the method is still widely used with the less important species that have not yet been heavily selected.

A variation of the pure-line selection method that dates back centuries is the selection of single- chance variants, mutations or “sports” in the original variety. A very large number of varieties that differ from the original strain in characteristics such as colour, lack of thorns or barbs, dwarfness, and disease resistance have originated in this fashion.

Hybridization

During the 20th century planned hybridization between carefully selected parents has become dominant in the breeding of self-pollinated species. The object of hybridization is to combine desirable genes found in two or more different varieties and to produce pure-breeding progeny superior in many respects to the parental types.

Genes, however, are always in the company of other genes in a collection called a genotype. The plant breeder’s problem is largely one of efficiently managing the enormous numbers of genotypes that occur in the generations following hybridization. As an example of the power of hybridization in creating variability, a cross between hypothetical wheat varieties differing by only 21 genes is capable of producing more than 10,000,000,000 different genotypes in the second generation. At spacing normally used by farmers, more than 50,000,000 acres would be required to grow a population large enough to permit every genotype to occur in its expected frequency. While the great majority of these second generation genotypes are hybrid (heterozygous) for one or more traits, it is statistically possible that 2,097,152 different pure-breeding (homozygous) genotypes can occur, each potentially a new pure-line variety. These numbers illustrate the importance of efficient techniques in managing hybrid populations, for which purpose the pedigree procedure is most widely used.

Pedigree breeding starts with the crossing of two genotypes, each of which have one or more desirable characters lacked by the other. If the two original parents do not provide all of the desired characters, a third parent can be included by crossing it to one of the hybrid progeny of the first generation (F1). In the pedigree method superior types are selected in successive generations, and a record is maintained of parent–progeny relationships.

The F2 generation (progeny of the crossing of two F1 individuals) affords the first opportunity for selection in pedigree programs. In this generation the emphasis is on the elimination of individuals carrying undesirable major genes. In the succeeding generations the hybrid condition gives way to pure breeding as a result of natural self-pollination, and families derived from different F2 plants begin to display their unique character. Usually one or two superior plants are selected within each superior family in these generations. By the F5 generation the pure-breeding condition (homozygosity) is extensive, and emphasis shifts almost entirely to selection between families. The pedigree record is useful in making these eliminations. At this stage each selected family is usually harvested in mass to obtain the larger amounts of seed needed to evaluate families for quantitative characters. This evaluation is usually carried out in plots grown under conditions that simulate commercial planting practice as closely as possible. When the number of families has been reduced to manageable proportions by visual selection, usually by the F7 or F8 generation, precise evaluation for performance and quality begins. The final evaluation of promising strains involves (1) observation, usually in a number of years and locations, to detect weaknesses that may not have appeared previously; (2) precise yield testing; and (3) quality testing. Many plant breeders test for five years at five representative locations before releasing a new variety for commercial production.

The bulk-population method of breeding differs from the pedigree method primarily in the handling of generations following hybridization. The F2 generation is sown at normal commercial planting rates in a large plot. At maturity the crop is harvested in mass, and the seeds are used to establish the next generation in a similar plot. No record of ancestry is kept. During the period of bulk propagation tends to eliminate plants having poor survival value. Two types of artificial selection also are often applied: (1) destruction of plants that carry undesirable major genes and (2) mass techniques such as harvesting when only part of the seeds are mature to select for early maturing plants or the use of screens to select for increased seed size. Single plant selections are then made and evaluated in the same way as in the pedigree method of breeding. The chief advantage of the bulk population method is that it allows the breeder to handle very large numbers of individuals inexpensively.

Often an outstanding variety can be improved by transferring to it some specific desirable character that it lacks. This can be accomplished by first crossing a plant of the superior variety to a plant of the donor variety, which carries the trait in question, and then mating the progeny back to a plant having the genotype of the superior parent. This process is called backcrossing. After five or six backcrosses the progeny will be hybrid for the character being transferred but like the superior parent for all other genes. Selfing the last backcross generation, coupled with selection, will give some progeny pure breeding for the genes being transferred. The advantages of the backcross method are its rapidity, the small number of plants required, and the predictability of the outcome. A serious disadvantage is that the procedure diminishes the occurrence of chance combinations of genes, which sometimes leads to striking improvements in performance.

Hybrid varieties The development of hybrid varieties differs from hybridization. The of crosses between different genotypes is often much more vigorous than its parents. This hybrid vigour, or heterosis, can be manifested in many ways, including increased rate of growth, greater uniformity, earlier flowering, and increased yield, the last being of greatest importance in agriculture.

CROSS POLLINATED CROPS

The most important methods of breeding cross-pollinated species are (1) mass selection; (2) development of hybrid varieties; and (3) development of synthetic varieties. Since cross-pollinated species are naturally hybrid (heterozygous) for many traits and lose vigour as they become (homozygous), a goal of each of these breeding methods is to preserve or restore heterozygosity. Mass selection

Mass selection in cross-pollinated species takes the same form as in self-pollinated species; i.e., a large number of superior appearing plants are selected and harvested in bulk and the seed used to produce the next generation. Mass selection has proved to be very effective in improving qualitative characters, and, applied over many generations, it is also capable of improving quantitative characters, including yield, despite the low heritability of such characters. Mass selection has long been a major method of breeding cross-pollinated species, especially in the economically less important species.

Hybrid varieties

The outstanding example of the exploitation of hybrid vigour through the use of F1 hybrid varieties has been with corn (maize). The production of a hybrid corn variety involves three steps: (1) the selection of superior plants; (2) selfing for several generations to produce a series of inbred lines, which although different from each other are each pure-breeding and highly uniform; and (3) crossing selected inbred lines. During the inbreeding process the vigour of the lines decreases drastically, usually to less than half that of field-pollinated varieties. Vigour is restored, however, when any two unrelated inbred lines are crossed, and in some cases the F1 hybrids between inbred lines are much superior to open-pollinated varieties. An important consequence of the homozygosity of the inbred lines is that the hybrid between any two inbreds will always be the same. Once the inbreds that give the best hybrids have been identified, any desired amount of hybrid seed can be produced.

Pollination in corn (maize) is by wind, which blows pollen from the tassels to the styles (silks) that protrude from the tops of the ears. Thus controlled cross-pollination on a field scale can be accomplished economically by interplanting two or three rows of the seed parent inbred with one row of the pollinator inbred and detasselling the former before it sheds pollen. In practice most hybrid corn is produced from “double crosses,” in which four inbred lines are first crossed in pairs (A × B and C × D) and then the two F1 hybrids are crossed again (A × B) × (C × D). The double-cross procedure has the advantage that the commercial F1 seed is produced on the highly productive single cross A × B rather than on a poor-yielding inbred, thus reducing seed costs. In recent years cytoplasmic male sterility, described earlier, has been used to eliminate detasselling of the seed parent, thus providing further economies in producing hybrid seed.

Much of the hybrid vigour exhibited by F1 hybrid varieties is lost in the next generation. Consequently, seed from hybrid varieties is not used for planting stock but the farmer purchases new seed each year from seed companies.

Perhaps no other development in the biological sciences has had greater impact on increasing the quantity of food supplies available to the world’s population than has the development of hybrid corn (maize). Hybrid varieties in other crops, made possible through the use of male sterility, have also been dramatically successful and it seems likely that use of hybrid varieties will continue to expand in the future.

Synthetic varieties

A synthetic variety is developed by intercrossing a number of genotypes of known superior combining ability—i.e.,genotypes that are known to give superior hybrid performance when crossed in all combinations. (By contrast, a variety developed by mass selection is made up of genotypes bulked together without having undergone preliminary testing to determine their performance in hybrid combination.) Synthetic varieties are known for their hybrid vigour and for their ability to produce usable seed for succeeding seasons. Because of these advantages, synthetic varieties have become increasingly favoured in the growing of many species, such as the forage crops, in which expense prohibits the development or use of hybrid varieties.

Source:http://www.britannica.com/EBchecked/topic/463294/plant-breeding/

MUTATION BREEDING

Physical Mutagens

Physical mutagens include various types of radiation, viz X-rays, gamma rays, alpha particles, beta particles, fast and thermal (slow) neutrons and ultra violet rays. A brief description of these mutagens is presented below: Commonly used physical mutagens (radiations), their properties and mode of action.

Type of Radiation Main properties

X – rays S.I., penetrating and non-particulate Gamma rays S.I., very penetrating and Non-particulate Alpha Particles D.I., particulate, less penetrating and positively charged. Beta Rays Particles S.I., particulate, more penetrating than alpha particles and negatively charged. Fast and Thermal D.I., particulate, neutral particles, highly penetrating. Neutrons 6. Ultra Violet Rays Non-ionizing, low penetrating

Note: particulate refers to particle emitting property DI = Densely ionizing, SI = Sparsely ionizing.

X-rays

X-rays were first discovered by Roentgen in 1895. The wavelengths of X-rays vary from 10-11 to 10-7. They are sparsely ionizing and highly penetrating. They are generated in X-rays machines. X- rays can break chromosomes and produce all types of mutations in nucleotides, viz. addition, deletion, inversion, transposition, transitions and transversions. X-rays were first used by Muller in 1927 for induction of mutations in Drosophila. In plants, Stadler in 1928 first used X-rays for induction of mutations in barley.

Gamma rays

Gamma rays have shorter wave length than X-rays and are more penetrating than gamma rays. They are generated from radioactive decay of some elements like 14C, 60Co, radium etc. Of these, cobalt 60 is commonly used for the production of Gamma rays. Gamma rays cause chromosomal and gene mutations like X-rays. CHEMICAL MUTAGENS

Procedure for chemical mutagenesis

The chemical mutagens can be divided into four groups, viz. 1) alkylating agents, 2) base analogues, 3) acridine dyes, and 4) others. A brief description of some commonly used chemicals of these groups is presented below.

Some commonly used chemical mutagens and their mode of action

Group of mutagen Name of chemical Mode of action

1. Alkylating Agents Ethyl methane Sulphonate AT GC Transitions Methyl Methane Sulphonate Transitions Ethyl Ethane Sulphonate GC AT Transitions Ethylene Imines Transitions

5 Bromo Uracil AT GC Transitions 1. Base Analogues 2 Amino purine AT GC Transitions 1. Acridine Dyes Acriflavin, Proflavin Deletion, addition and frame shifts.

AT GC Transitions Nitrous Acid GC AT Transitions 1. Others Hydroxylamine Transitions Sodium Azide

The speed of hydrolysis of the chemical mutagens is usually measured by the half life of the chemicals. Half life is the time required for disappearance of the half of the initial amount of active reaction agent.

In the case of DES the mutagenic solution should be changed at every half an hour to get good results. Half life is the function of temperature and pH for a particular compound.

One should be extremely careful in handling alkylating agents since most of them are carcinogenic. Especially for ethylene imine, it should be handled under aerated conditions. EMS though not dangerous, it should not be pipetted out by mouth. Besides the alkylating agents, we are also having chemical mutagens like, Base analogues, Acridine dyes, Antibiotics and other miscellaneous chemicals.

Treatment of seeds with mutagenic chemicals:

Materials required:- conical flask, beaker, pipette, glass rods, measuring cylinder, stop watch, distilled water and phosphate buffer.

Method: - Mutagenic chemical is diluted to the required concentration by using distilled water. To prepare the molar concentration of DES, the method is

Molecular weight x a.i. (purity percentage) Specific gravity (active ingredient)

154 100 Eg. DES = ---- x ----- = 131 CC.

1. 18 ...... 99

131 CC dissolved in one litre will give 1 molar solution.

Seeds have to be soaked in the distilled water for different hours depending upon the seeds, to initiate biochemical reactions. The chemical action is found to be affected by the frequency and spectrum of mutagen depending upon the stage of cell division, during the process of germination. If the chemical treatment is synchronized with DNA synthesis stage (G1, S and G2) then we can get better results.

The presoaked seeds are taken in a flask and chemical is added. Usually the quantity of the chemical is ten times the volume of seeds. Intermittent shaking should be given to ensure uniform exposure of the chemicals. The chemical should be drained after the treatment time is over. The seeds should be washed thoroughly in running tap water, immediately for not less than 30 minutes. After washing, the seeds should be dried in between the filter paper folds. Seeds are to be arranged in germination tray with equal spacing. Trays are kept in a controlled environment of temperature and humity. Periodical observation on germination upto 10-15 days is needed. From the germination percentage, we can assess the LD50 dose.

Plant Genetic Resources

The sum total of hereditary material i.e. all the alleles of various genes, present in a crop species and its wild relatives is referred to as germplasm. This is also known as genetic resources or gene pool or genetic stock. Important features of plant genetic resources are given below.

 Genetic pool represents the entire genetic variability or diversity available in a crop species.  Germplasm consists of land races, modern , obsolete cultivars, breeding stocks, wild forms and wild species of cultivated crops.  Germplasm includes both cultivated and wild species and relatives of crop plants.  Germplasm is collected from centres of diversity, gene banks, gene sanctuaries, farmer’s fields, markers and seed companies.  Germplams is the basic material for launching a crop improvement programme.  Germplasm may be indigenous (collected within country) or exotic (collected from foreign countries)

Germplasm Conservation

Conservation refers to protection of genetic diversity of crop plants from genetic erosion. There are two important methods of germpalsm conservation or preservation. i) In-situ conservation and ex situ conservation. These are described below. i) In - situ conservation:

Conservation of germplasm under natural conditions is referred to as in situ conservation. This is achieved by protecting the area from – human interference, such an area is often called natural park, biosphere reserve or gene sanctuary. NBPGR, New Delhi, established gene sanctuaries in Meghalaya for citrus, north Eastern regions for musa, citrus, oryza and saccharum. Gene sanctuaries offer the following advantage.

Merits: In this method of conservation, the wild species and the compete natural or seminatural ecosystems are preserved together.

Demerits:

 Each protected area will cover only very small portion of total diversity of a crop species, hence several areas will have to be conserved for a single species.  The management of such areas also poses several problems.  This is a costly method of germplasm conservation. ii) Ex - situ conservation:

It refers to preservation of germplasm in gene banks. This is the most practical method of germplasm conservation. This method has following advantages.

 It is possible to preserve entire genetic diversity of a crop species at one place.  Handling of germplasm is also easy.  This is a cheap method of germplams conservation.

This type of conservation can be achieved in the following 5 ways.

1) Seed banks:

Germplam is stored as seeds of various genotypes. Seed conservation is quite easy, relatively safe and needs minimum space. Seeds are classified, on the basis of their storability into two major groups.

1) Orthodox and 2) Recalcitrant

Orthodox seeds: Seeds which can be dried to low moisture content and stored at low temperature without losing their viability for long periods of time is known as orthodox seeds. (eg.) Seeds of corn, wheat, rice, carrot, papaya, pepper, chickpea, cotton, sunflower. Recalcitrant: Seeds which show very drastic loss in viability with a decrease in moisture content below 12 to 13% are known as recalcitrant seeds. (e.g) citrus, cocoa, coffee, rubber, oilpalm, mango, jack fruit etc.

Seed storage: Based on duration of storage, seed bank collects are classified into three groups. (1) Base collections. (2) Active collections and (3) Working collection.

Base collections: Seeds can be conserved under long term (50 to 100 years), at about -20OC with 5% moisture content. They are disturbed only for regeneration.

Active collection: Seeds are stored at 0OC temperature and the seed moisture is between 5 and 8%. The storage is for medium duration, i.e., 10-15 years. These collections are used for evaluation, multiplication, and distribution of the accessions.

Working collections: Seeds are stored for 3-5 years at 5-10OC and the usually contain about 10% moisture. Such materials are regularly used in crop improvement programmes.

2. Plant Bank: ( Field or plant bank )is an orchard or a field in which accessions of fruit trees or vegetatively propagated crops are grown and maintained.

Limitations:

1. Require large areas 2. Expensive to establish and maintain 3. Prone to damage from disease and insect attacks 4. Man – made 5. Natural disasters 6. Human errors in handling

Shoot tip banks: Germplasm is conserved as slow growth cultures of shoot-tips and node segments. Conservation of genetic stocks by meristem cultures has several advantages as given below.

 Each genotype can be conserved indefinitely free from virus or other pathogens.  It is advantageous for vegetatively propagated crops like potato, sweet potato, cassava etc., because seed production in these crops is poor  Vegetatively propagated material can be saved from natural disasters or pathogen attack.  Long regeneration cycle can be envisaged from meristem cultures.  Regeneration of meristerms is extremely easy.  Plant species having recalcitrant seeds can be easily conserved by meristem cultures.

Cell and organ banks: A germplasm collection based on cryopreserved (at – 196OC in liquid nitrogen) embryogenic cell cultures, somatic/ zygotic embryos they be called cell and organ bank.

DNA banks: In these banks, DNA segments from the genomes of germplasm accessions are maintained and conserved.

Germplam evaluation

Evaluation refers to screening of gemplasms in respect of morphological, genetical, economic, biochemical, physiological, pathological and entomological attributes. Evaluation of germplasm is essential from following angles.

 To identify gene sources for resistance to biotic and abiotic stresses, earliness, dwarfness, productivity and quality characters.  To classify the germplasm into various groups  To get a clear pictures about the significance of individual germplasm line.

IPGRI, Rome has developed model list of descriptors (= characters) for which germplasm accessions of various crops should be evaluated. The evaluation of germplasm is done in three different places viz., (1) in the field (2) in green house a) 3) in the laboratory.

Germplasm cataloguing, Data storage and Retrieval.

Each germplasm accession is given an accession number. This number is pre fixed in India, with either IC (Indigenous collection), EC (exotic collection) or IW (Indigenous wild). Information on the species and variety names, place of origin, adaptation and on its various feature or descriptors is also recorded in the germplasm maintenance records. Catalogues of the germplasm collection for various crops are published by the gene banks. The amount of data recorded during evaluation is huge. Its compilation, storage and retrieval is now done using special computer programmes.

National Bureau of Plant Genetic Resources (NBPGR)

NBPGR establishment in 1976 is the nodal organisation in India for planning, conducting, promoting, coordinating and lending all activities concerning plant.

 Collection  Introduction  Exchange  Evaluation  Documentation  Safe conservation  Sustainable management of germplasm

Vegetable Crop Responsibilities and Germplasm Activities at NBPGR

The vegetable crop germplasm programme broadly includes the following vegetable crops for evaluation, documentation and maintenance of active collections besides their long term storage:

A. Solanaaceous : Brinjal, tomato, chillies B. Cucurbitaceous Vegetables : Pumpkin, melons, gourds and cucumber C. Leguminous vegetables : Cowpean, pea, lablab bean, winged bean, faba bean, French bean D. Bulb crops : Garlic, onion E. Root vegetables : Radish, carrot, turnip F. Okra : - G. Miscellaneous vegetables : Cole crops, Chinese cabbage, spinach beet, spinach

The quantum of variability available and of diversity of various vegetable crops shows that India is one of the important centres/regions of variability of vegetable crops. The centre of origin/diversity of various vegetable crops reveals that a number of vegetable crops of economic importance and their wild relatives originated in this region. These genetic resources possess genes for wide adaptability, high yield potential including resistance/tolerance to biotic and abiotic stresses. The Indian sub- continent, thus holds prominence as one of the twelve regions of variability in crop plants in global perspective.

Gene banks for various crops in India

Institutes Crops Central Institute for Cotton Research, Nagpur Cotton Central Plantation crops Research Institute, Kasargod Plantation crop Central Potato Research Institute, Simla Potato Central tobacco research Institute, Rajahmundry Tobacco Central tuber crops research Institute,Tuber crops other than potato Thiruvananthapuram Central Rice Research Institute, Cuttack Rice Directorate of Oilseeds research, Hyderabad Oilseeds Directorate of Wheat Research, Karnal Wheat Indian Agricultural Research Institute, New Delhi Maize Indian Grassland and Fodder Research Institute, Jhansi Forge and fodder crops National research centre for sorghum, Hyderabad Sorghum International Crops Research Institute for Semi-AridGroundnut, Pearl millet, Sorghum, Tropics Pigeon pea and Bengal gram

List of important International Institutes conserving germplasm

Name Institute Activity IRRI International Rice Research Institute, LosTropical rice Banos, Philippines Rice collection: 42,000 CIMMYT Centre International de-Mejoramients deMaize and wheat (Triticale, maize Trigo, El Baton, Mexico barely, sorghum) Maize collection – 8000 CIAT Center International de-agriculturalCassava and beans, (also maize Tropical Palmira, Columbia and rice) in collobaration with CIMMYT and IRRI IITA International Institute of TropicalGrain legumes, roots, and tubers, Agriculture, Ibadan, Nigeria. farming systems. CIP Centre International de-papa-Lima. Peru Potatoes ICRISAT International Crops Research Institute, forSorghum, Groundnut, Cumbu, Semi-Arid Tropics, Hyderabad, India Bengalgram, Redgram. WARDA West African Rice DevelopmentRegional Cooperative Rice Association, Monrovia, Liberia Research in Collaboration with IITA and IRRI IPGRI International Plant Genetic ResearchGenetic conservation. Institute, Rome Italy AVRDC The Asian Vegetable Research andTomato, Onion, Peppers Chinese Development Centre, Taiwan cabbage.

MODE OF REPRODUCTION

Knowledge of the mode of reproduction and pollination is essential for a plant breeder, because these aspects help in deciding the breeding procedures to be used for the genetic improvement of a crop species. Choice of breeding procedure depends on the mode of reproduction and pollination of a crop species.

Reproduction refers to the process by which living organisms give rise to the offspring of similar kind (species). In crop plants, the mode of reproduction is of two types: viz. 1) sexual reproduction and 2) asexual reproduction

I. Sexual reproduction

Multiplication of plants through embryos which have developed by fusion of male and female gametes is known as sexual reproduction. All the seed propagating species belong to this group.

Sporogenesis

Production of microspores and megaspores is known as sporogenesis. In anthers, microspores are formed through microsporogensis and in ovules, the megaspores are formed through megasporogenesis. Microsporogenesis

The sporophytic cells in the pollen sacs of anther which undergo meiotic division to form haploid i.e., microspores are called microspore (MMC) or pollen mother cell (PMC) and the process is called microsporogenesis. Each PMC produce four microspores and each microspore after thickening of the wall transforms into pollen grain.

Megasporogenesis

A single sporophytic cell inside the ovule, which undergo meiotic division to form haploid megaspore, is called megaspore mother cell (MMC) and the process is called megasporogenesis. Each MMC produces four megaspores out of which three degenerate resulting in a single functional megaspore.

Gametogenesis

The production of male and female gametes in the microspores and megaspores is known as gametogenesis.

Microgametogenesis

This is nothing but the production of male gametes or sperm. On maturation of the pollen, the microspore nucleus divides mitotically to produce a generative and a vegetative or tube nucleus. The pollen is generally released in this binucleate stage. The reach of pollen over the stigma is called pollination. After the pollination, the pollen germinates. The pollen tube enters the stigma and travels down the style. The generative nucleus at this phase undergoes another mitotic division to produce two male gametes or sperm nuclei. The pollen along with the pollen tube possessing a pair of sperm nuclei is called microgametophyte. The pollen tube enters the embryo sac through micropyle and discharges the two sperm nuclei.

Megagametogenesis

The nucleus of the functional megaspore undergoes three mitotic divisions to produce eight or more nuclei. The exact number of nuclei and their arrangement varies from one species to another. The megaspore nucleus divides thrice to produce eight nuclei. Three of these nuclei move to one pole and produce a central egg cell and two synergid cells on either side. Another three nuclei migrate to the opposite pole to develop into three antipodal cells. .

The two nuclei remaining in the center, the polar nuclei, fuse to form the secondary nucleus. The megaspore thus develops into a mature female gametophyte called megagametophyte or embryo sac. The development of embryo sac from a megaspore is known as megagametogeneis. The embryo sac generally contains one egg cell, two synergids with the apparent function of guiding the sperm nucleus towards the egg cell and three antipodals which forms the prothalamus cells and one diploid secondary nucleus.

Fertilization:

The fusion of one of the two sperms with the egg cell producing a diploid zygote is known as fertilization. The fusion of the remaining sperm with the secondary nucleus leading to the formation of a triploid primary endosperm nucleus is termed as triple fusion. The primary endosperm nucleus after several mitotic divisions develops into mature endosperm, which nourishes the developing embryo

II. Asexual reproduction

Multiplication of plants without the fusion of male and female gametes is known as asexual reproduction. Asexual reproduction can occur either by vegetative plant parts or by vegetative embryos which develop without sexual fusion (apomixis). Thus asexual reproduction is of two types: viz. a) vegetative reproduction and b) apomixis. Vegetative reproduction refers to multiplication of plants by means of various vegetative plant parts. Vegetative reproduction is again of two types: viz. i) natural vegetative reproduction and ii) artificial vegetative reproduction.

Natural vegetative reproduction

In nature, multiplication of certain plants occurs by underground stems, sub aerial stems, roots and bulbils. In some crop species, underground stems (a modified group of stems) give rise to new plants. Underground stems are of four types: viz. rhizome, tuber, corm and bulb. The examples of plants which reproduce by means of underground stems are given below:

Rhizome: Turmeric (Curcuma domestica), Ginger (Zingiber officinale) Tuber: Potato (Solanum tuberosum) Corm: Arvi (Colocasia esculenta), Bunda (C. antiquorum) Bulb: Garlic (Allium sativum), onion (A. cepa)

Rhizome: Turmeric Tuber: Potato Bulb: Onion

Sub aerial stems include runner, sucker, stolon, etc. These stems lead to vegetative reproduction in mint (Mentha sp) rose, strawberry, banana, etc. Bulbils are modified forms of flower. They develop into plants when fall on the ground. Bulbils are found in garlic. Artificial vegetative reproduction: Multiplication of plants by vegetative parts through artificial method is known as artificial vegetative reproduction. Such reproduction occurs by cuttings of stem and roots, and by layering and grafting. Examples of such reproduction are given below: Stem cuttings: Sugarcane (Saccharum sp.) grapes (Vitis vinifera), roses, etc. Root cuttings: Sweet potato, citrus, lemon, etc.

Layering and grafting are used in fruit and ornamental crops.

Apomixis

Apomixis refers to the development of seed without sexual fusion (fertilization). In apomixis embryo develops without fertilization. Thus apomixis is an asexual means of reproduction. Apomixis is found in many crop species. Reproduction in some species occurs only by apomixis. This apomixis is termed as obligate apomixis. But in some species sexual reproduction also occurs in addition to apomixis. Such apomixis is known as facultative apomixis.

There are four types of apomixis: viz.

1) parthenogenesis, 2) apogamy, 3) apospory and 4) adventive embryony.

1. Parthenogenesis. Parthenogenesis refers to development of embryo from the egg cell without fertilization.

2. Apogamy. The origin of embryo from either synergids or antipodal cells of the embryosac is called as apogamy.

3. Apospory. In apospory, first diploid cell of ovule lying outside the embryosac develops into another embryosac without reduction. The embryo then develops directly from the diploid egg cell without fertilization.

4. Adventive embryony. The development of embryo directly from the diploid cells of ovule lying outside the embryosac belonging to either nucellus or integuments is referred to as adventive embryony.

MODE of Pollination

The process by which pollen grains are transferred from anthers to stigma is referred as pollination. Pollination is of two types: viz. 1) Autogamy or self pollination and 2) Allogamy or cross pollination.

I. Autogamy

Transfer of pollen grains from the anther to the stigma of same flower is known as autogamy or self pollination. Autogamy is the closest form of inbreeding. Autogamy leads to homozygosity. Such species develop homozygous balance and do not exhibit significant .

Mechanism promoting self-pollination 1. Bisexuality. Presence of male and female organs in the same flower is known as bisexuality. The presence of bisexual flowers is a must for self pollination. All the self pollinated plants have hermaphrodite flowers.

2. Homogamy. Maturation of anthers and stigma of a flower at the same time is called homogamy. As a rule, homogamy is essential for self-pollination.

3. Cleistogamy. When pollination and fertilization occur in unopened flower bud, it is known as cleistogamy. It ensures self pollination and prevents cross pollination. Cleistogamy has been reported in some varieties of wheat, barley, oats and several other grass species.

4. Chasmogamy. Opening of flowers only after the completion of pollination is known as chasmogamy. This also promotes self pollination and is found in crops like wheat, barley, rice and oats.

5. Position of Anthers. In some species, stigmas are surrounded by anthers in such a way that self pollination is ensured. Such situation is found in tomato and brinjal. In some legumes, the stamens and stigma are enclosed by the petals in such a way that self pollination is ensured. Examples are greengram, blackgram, soybean, chickpea and pea.

II. Allogamy

Transfer of pollen grains from the anther of one plant to the stigma of another plant is called allogamy or cross pollination. This is the common form of out-breeding. Allogamy leads to heterozygosity. Such species develop heterozygous balance and exhibit significant inbreeding depression on selfing. Mechanism promoting cross-pollination

1. Dicliny. It refers to unisexual flowers. This is of two types: viz. i) monoecy and ii) dioecy. When male and female flowers are separate but present in the same plants, it is known as monoecy. In some crops, the male and female flowers are present in the same inflorescence such as in mango, castor and banana. In some cases, they are on separate inflorescence as in maize. Other examples are cucurbits, grapes, strawberry, cassava and rubber. When staminate and pistillate flowers are present on different plants, it is called dioecy. It includes papaya, date palm, spinach, hemp and asparagus.

2. Dichogamy. (from the Greek dikho-apart and gamous-marriage) It refers to maturation of anthers and stigma of the same flowers at different times. Dichogamy promotes cross pollination even in the hermaphrodite species. Dichogamy is of two types: viz. i) protogyny and ii) protandry. When pistil matures before anthers, it is called protogyny such as in pearl millet. When anthers mature before pistil, it is known as protandry. It is found in maize, sugarbeet and several other species.

3. Heterostyly. When styles and filaments in a flower are of different lengths, it is called heterostyly. It promotes cross pollination, such as linseed.

4. Herkogamy. Hinderance to self-pollination due to some physical barriers such as presence of hyline membrane around the anther is known as herkogamy. Such membrane does not allow the dehiscence of pollen and prevents self-pollination such as in alfalfa.

5. Self incompatibility: The inability of fertile pollens to fertilize the same flower is referred to as self incompatibility. It prevents self-pollination and promotes cross pollination. Self incompatibility is found in several crop species likeBrassica, Radish, Nicotiana, and many grass species. It is of two types sporophytic and gametophytic.

6. Male sterility: In some species, the pollen grains are non functional. Such condition is known as male sterility. It prevents self-pollination and promotes cross pollination. It is of three types: viz. genetic, cytoplasmic and cytoplasmic genetic. It is a useful tool in hybrid seed production.

Study of floral biology and aforesaid mechanisms is essential for determining the mode of pollination of various crop species. Moreover, if selfing has adverse effects on seed setting and general vigour, it indicates that the species is cross pollinated. If selfing does not have any adverse effect on these characters, it suggests that the species is self-pollinated.

The percentage of cross pollination can be determined by growing a seed mixture of two different varieties together. The two varieties should have marker characters say green and pigmented plants. The seeds are harvested from the recessive (green) variety and grown next year in separate field. The proportion of pigmented plants in green variety will indicate the percentage of or cross pollination.

Significance of pollination

The mode of pollination plays an important role in plant breeding. It has impact on five important aspects : viz. 1) gene action, 2) genetic constitution, 3) adaptability, 4) genetic purity and 5) transfer of genes.

Classification of crop plants based on mode of pollination and mode of reproduction

Mode of pollination andExamples of crop plants reproduction A. Autogamous Species 1. Seed Propagated Rice, Wheat, Barley, Oats, Chickpea, Pea, Cowpea, Lentil, Green gram, Black gram, Soybean, Common bean, Moth bean, Linseed, Sesame, Khesari, Sunhemp, Chillies, Brinjal, Tomato, Okra, Peanut, etc. 2. Vegetatively Propagated Potato B. Allogamous Species 1. Seed Propagated Corn, Pearlmillet, Rye, Alfalfa, Radish, Cabbage, Sunflower, Sugarbeet, Castor, Red clover, White clover, Safflower, Spinach, Onion, Garlic, Turnip, Squash, Muskmelon, Watermelon, Cucumber, Pumpkin, Kenaf, Oilpalm, Carrot, Coconut, Papaya, etc. 2. Vegetatively propagated Sugarcane, Coffee, Cocoa, Tea, Apple, Pears, Peaches, Cherries, grapes, Almond Strawberries, Pine apple, Banana, Cashew, Irish, Cassava, Taro, Rubber, etc. C. Often Allogamous Species Sorghum, Cotton, Triticale, Pigeonpea, Tobacco.

Selfing and Crossing Techniques: elfing and crossing are the essential procedures in crop improvement process. The exact procedures used to ensure self or cross-pollination of specific plants will depend on the floral structure and normal manner of pollination. Generally effecting cross-pollination in a strictly self-pollinating species is more difficult than vice-versa because for instance preventing self-pollination occurring inside the unopened flowers is cumbersome.

Selfing

In the selfing of cross-pollinated species, it is essential that the flower are bagged or otherwise protected to prevent natural cross-pollination. Selfing and crossing are essential in crop breeding. It is important that the breeder, master these techniques in order to manipulate the pollination according to his needs. The exact procedure that he may use to ensure self or cross pollination of specific plants will depend on the particular species with which he is working. The structure of the flowers in the species determine manner of pollination. For these reasons, the breeder should acquaint himself with the flowering habit of the crop.

In the case of wheat, rice, barely, groundnut etc., the plant is permitted to have self pollination and the seeds are harvested. It is necessary to know the mode of pollination. If the extent of natural cross pollination is more, then the flowers should be protected by bagging. This will prevent the foreign pollen to reach the stigma. Seed set is frequently reduced in ear heads enclosed in bags because of excessive temperature and humidity inside the bags. In crops like cotton which have larger flowers the petals may fold down the sexual organs and fasten, there by pollen and pollen carrying insects may be excluded.

In certain legumes which are almost insect pollinated, the plants may be caged to prevent the insect pollination. In maize, a paper bag is placed over the tassel to collect pollen and the cob is bagged to protect from foreign pollen. The pollen collected from the tassel is transferred to the cob.

Emasculation Removal of stamens or anthers or killing the pollen of a flower without the female reproductive organ is known as emasculation. In bisexual flowers, emasculation is essential to prevent of self- pollination. In monoecious plants, male flowers are removed. (castor, coconut) or male inflorescence is removed (maize). In species with large flowers e.g. (cotton, pulses) hand emasculation is accurate and it is adequate.

Methods of Emasculation

1. Hand Emasculation

In species with large flowers, removal of anthers is possible with the help of forceps. It is done before anther dehiscence. It is generally done between 4 and 6 PM one day before anthers dehisce. It is always desirable to remove other young flowers located close to the emasculated flower to avoid confusion. The corolla of the selected flower is opened with the help of forceps and the anthers are carefully removed with the help of forceps. Sometimes corolla may be totally removed along with epipetalous stamens e.g. gingelly.

In cereals, one third of the empty glumes will be clipped off with scissors to expose anthers. In wheat and oats, the florets are retained after removing the anthers without damaging the spikelets. In all cases, gynoecium should not be injured. An efficient emasculation technique should prevent self pollination and produce high percentage of seed set on cross pollination.

2. Suction Method

It is useful in species with small flowers. Emasculation is done in the morning immediately after the flowers open. A thin rubber or a glass tube attached to a suction hose is used to suck the anthers from the flowers. The amount of suction used is very important which should be sufficient to suck the pollen and anthers but not gynoecium. In this method considerable self-pollination, upto 10% is like to occur. Washing the stigma with a jet of water may help in reducing self-pollination, However self pollination can not be eliminated in this method.

3. Hot Water Treatment

Pollen grains are more sensitive than female reproductive organs to both genetic and environmental factors. In case of hot water emasculation, the temperature of water and duration of treatment vary from crop to crop. It is determined for every species. For sorghum 42-48OC for 10 minutes is found to be suitable. In the case of rice, 10 minutes treatments with 40-44OC is adequate. Treatment is given before the anthers dehiscence and prior to the opening of the flower. Hot water is generally carried in thermos flask and whole inflorescence is immersed in hot water.

4. Alcohol Treatment

It is not commonly used. The method consists of immersing the inflorescence in alcohol of suitable concentration for a brief period followed by rinsing with water. In Lucerne the inflorescence immersed in 57% alcohol for10 second was highly effective. It is better method of emasculation than suction method. 5.Cold Treatment

Cold treatment like hot water treatment kills the pollen grains without damaging gynoecium. In the case of rice, treatment with cold water 0.6OC kills the pollen grains without affecting the gynoecium. This is less effective than hot water treatment.

6. Genetic Emasculation

Genetic/ cytoplasmic male sterility may be used to eliminate the process of emasculation. This is useful in the commercial production of hybrids in maize, sorghum pearlmillet, onion, cotton, and rice, etc.,In many species of self-incompatible cases, also emasculation is not necessary, because self-fertilization will not take place. Protogyny will also facilitate crossing without emasculation (e.g.) Cumbu.

7. Use of Gametocide

Also known as chemical hybridizing agents (CHA) chemicals which selectively kills the male gamete without affecting the female gamete. eg. Ethrel, Sodium methyl arsenate, Zinc methyl arsenate in rice, Maleic hydrazide for cotton and whe

Bagging

Immediately after emasculation the flower or inflorescence enclosed with suitable bags of appropriate size to prevent random cross-pollination.

Pollination

The pollen grains collected from a desired male parent should be transferred to the emasculated flower. This is normally done in the morning hours during anthesis. The flowers are bagged immediately after artificial crossing.

Tagging

The flowers are tagged just after bagging. They are attached to the inflorescence or to the flower with the help of a thread. The following may be recorded on the tag with pencil.

1. Date of emasculation 2. Date of pollination 3. Parentage 4. No. of flowers emasculate Proc. Indian Acad. Sci. (Plant Sci.), Vol. 93, No. 3, .Iuly 1984, pp. 401-412 Printed in India.

Forest tree improvement in India

S KEDHARNATH Kerala Forcst Rcsearch Institutr Pr 680 653, India Abstract. Forest tree breeding is relativelya young science. Even so, there is good evidenceof its potentiality for increasing forest productivityand quality of the forest produce. The basic scheme for forest tree improvement involves selection of superior parent trees, assembling them as ciones in seed orchards in special designs to promote maximum cross pollination among the different dones and reduce inbreeding. Interprovenance and interspecific hyb¡ are also resorted to in special situations. Forest tree improvementwork through selection and breeding has been in progressin India for the last nearly two decades. Someof the achievements and strategies used are briefly reviewed. Keywords. Forest tree breeding; seed orchards; ; eucalypts; pines; red sanders; semul; breeding strategies; interprovenance; inter-specific hyb¡

1. Introduction

Forest tree breeding per se is comparativcly a young science which had its recognizable start about rice decades back. Sweden was the pion•cr in this ventur• and otbers followed suit slowly. Today every country, big of small, developed of developing, has active programmes on forest tree breeding. In some of the developr countries wood- based industries hace invcsted money in such programmes with great expectations. Fortunately, there is sufficient good evidence today which indicates that such investments ate sound and can pay ¡ dividends. In fact in reccnt y•ars the research on the economics of tree improvement programmes has shifted its emphasis from programme justification to programme optimization. Intensive management through artificial regeneration and establishment of plan- tations of what may be termed as 'man-made forests' are increasingly being resorted to in many countries. This naturally offers a good opportunity for not only better management of the conditions under which the trer are to grow but also of choosing appropriate genotypes which will not only have the capacity to exploit to the best advantagc the environment provided but which can often be tailored to meet the specific needs. Sinc• the arca of potential production is often large even small improvcments in productivity may be very significant at the national levcl both in temas of social benefits and production of raw material. In this papr the general strategy that is followed in forest tree improvemcnt and the work currently in progress in India in this fascinating fi•ld ate b¡ reviewed.

2. Tree improvement strategy

2.1 Population improvement Exploitation ofavailable natural genetic variability within the species is the first step in all selcction and breeding work.

4O1 l'-- 25 402 S Kedharnath

A characte¡ feature ofall living organisms is the immense natural va¡ they exhibit for various characters in most populations. Broadly three types ofvariation may be recognised (i) random variation from tree-to-tree on the same site, (ii) va¡ in the average ofcertain characters ofall the trees in one locality of site when compared to the average of all trees in another locality (sometimes called local variation) and (iii) average vafiations in trees from widely different parts of the species range (often referred to as geographical variation or racial variation). In a regeneration programme it is essential first to identify the best adapted and productive seed origin of provenance for the species concerned. The thumb rule in forestry is "use seed from the local source until some other source or provenance has been proven superior to the local one". Superiority of new seed sources should be assessed by well laid out provenance trials. Having identified the ¡ provenance one can then exploit the tree-to-tree variation in economic traits for selection of superior individual trees. This is known as 'plus tree' selection. Plus trees may be defined as outstanding individuals occurring in natural stands or in even aged plantations combining in themselves a number of desirable features. As is to be expected such trees occur in low frequency and so may appear hard to find. But they do exist. These trees forro the foundation for tree improvement by selection. Kedharnath (1982) has briefly reviewed the different methods employed in plus tree selection. Plus trees which ate progeny tested and approved as good are called 'elite trees'. The number and type of characters used as selection criteria vary with the species but some of the most common characters used for selection ate good growth vigour, superior height growth, superior diameter growth, good pruning ability, straight cylindrical bole, narrow compact crown, resistance to important diseases and insect pests. The selected plus trees are then assembled as dones in special planting sites and they are called seed orchards. This is meant for mass producing quickly regular crops of genetically improved seeds for use in raising new plantations. These orchards require special management practices to enhance flower and seed production. Also, in the orchard the ramets of the different ciones ate so planted that there is very little inbreeding and maximum cross breeding between ramets of different ciones in various combinations are favoured. Special planting designs are used for this. Usually grafting is resorted to using scions collected from the top one third of the flowering crown of the plus trees. This increases the probability ofearly flowering in the grafted plants. In some cases rooted cuttings can also be employed ir graft incompatibility is a constraint. Seed orchard approach of mass producing genetically improved seeds in forest trees is so far the best publicised and most widely practised method in all the countries. This is most suitable for tree species that are normally cross pollinated. Also, to keep the genetic base sufficiently broad at least 20 ciones should be used in an orchard. Theoretically there is no upper limit to the number of ciones that can be used in an orchard. But the orchard should be large enough in arca so that various possible cross combinations among the ciones used could be realised.

2.2 Exploitation of hybrid viffour or heterosis Synthesising F 1 hybrids between selected provenances of a given species could be resorted to if some specific combination manifest hybrid vigour. Mass production of such F~ seeds could be achieved through a seed orchard programme where selected dones of two provenances may- be planted. The same approach can be followed for Forest tree improvement in India 403

obtaining F 1 hybrids between two selected species if there is primafacie evidence of hybrid vigour.

3. Work on tree improvement in India by selection and breeding

Organised work on breeding of forest trees was started in India in 1960 at the Forest Research Institute, Dehradun in some selected tree species of economic importance. The detailed programme of work to be initiated in forest genetics and tree breeding was presented at the Tenth Silvicultural Conference held at Dehradun by Kedharnath and Raizada (1961). The Forest Genetics Branch ofForest Research Institute, Dehradun in collaboration with the State Forest Departments have been active since 1960 in carrying out this programme ofwork and good progress has been registered in the case of teak (Tectona grandis L.f.) Chirpine (Pinus roxburgh¡ Sarg.), Semul (Bombax ceiba L.) and in some species of eucalypts. The Kerala Forest Research Institute, Peechi (Kerala), the Department ofForestry, H P Krishi Vishwa Vidyalaya and the Faculty of Forestry of Tamil Nadu Agricultural University huye been making very useful contributions in this field of work. Some of the important contributions in the area ate briefly reviewed below.

3.1 Teak (Tectona #randis L.f.) Teak is one of the most durable and valuable timber species and belongs to the family Verbenaceae. It has 36 us its somatic chromosome number (Kedharnath and Raizada 1961). It is native to the Indo-Malayan region and occurs naturally in some parts of India while in other parts it has been successfully introduced. Presently teak is being raised in plantations on a large scale. Approximately one lakh hectares ate being planted annually. The objective of tree improvement programme in teak has been to produce by selection and breeding superior stem forro, superior tate of growth in height and diameter, freedom from fluting, buttressing and epicgrmic branches, resistance to leaf skeletoniser (Eutectona machaeralis syn. Pyrausta machaeralis) and leal defoliator (Hyblaea purea). There is general evidence ofinherent variation in this species for all the characteristics stated above (Kedharnath and Matthews 1962). Work on the selection of plus trees in the species was started in the year 1960 and todate about 700 plus trees ate available for use in establishing clonal seed orchards. Simple budding and/or sometimes cleft grafting technique standardised for this species by Rawat and Kedharnath (1968) has been used for clonal seed orchards and germ plasm banks that huye been established in a number of states. The first experimental clonal seed orehard in this species was established at New Forest, Dehradun, Studies carried out on early growth performance of 20 dones revealed considerable variation between ciones (Kedharnath et al 1970). These differences have persisted in later years also. Similarly observations recorded over the years on the relative resistance/suscept- ibility of the different dones established in the germ plasta bank and in the clonal seed orchard at New Forest, Dehradun to two leaf infecting fun~---Olivaea tectonae and Caldariomyces tectonae--under natural conditions of infection showed consistent reaction. Some were absolutely resistant, some were very susceptible while some were moderately resistant. These two diseases are not economically important. In respect of studies on variation in fibre length carried out using trees from a replicated provenance 404 S Kedharnath experiment revealed significant geographical and tree-to-tree variation in this trait (Kedharnath et al 1963). Testing ofsome of the dones ofT. grandis anda related species T. hamiltoniana under controlled conditions for variation in resistance to Eutectona machaeralis showed that there is significant variation among the ciones tested (Kedharnath and Pratap Singh 1975). Studies carried out on vegetative propagation such as rooting of cuttings, grafting, budding etc have shown that it is a favourable material for cloning. There has not been any indication of graft incompatibility in T. grandis. Furthermore, budding on naked stumps (with about 15 cm length of root and about 3 cm of stem)just above the collar region gives very good take. Customarily such budded stumps are planted in polypots and the new sprout emerges in 15-20 days. The best time for such budding appears to be March-Ap¡ In June-July they are transplanted in the seed orchard site. More recently attempts have been made for clonal multiplication through in vitro tissue culture techniques and very good success has been obtained by Gupta et al (1980) at the National Chemical Laboratory, Pune. The important aspect of the above work is that mature excised terminal buds from field grown trees about I00 years old have been induced to form multiple shoots on a defined medium. Individual shoots were later made to develop roots on a low salt medium, containing three auxins. The plants so obtained have been later transplanted in pots and finally in the field. This technique offers a good method for cloning. But this is yet to become popular with foresters for large scale adoption in clonal seed orchard establishment. AII the teak-growing states are establishing clonal seed orchards for teak. It has been estimated that ifone plants 156 gratis per hectare at 8 x 8 m espacement 1280 ha of seed orchards will be needed. Roughly 2 grafted plants will suffice to give enough seeds (3 kg each) to planta hectare. About 800 ha of seed orchards have been established so lar. Progeny testing of the plus trees is ah essential step to know the breeding value of the plus trees. Open-pollinated seeds from the plus trees have been collected and used in some states like Tamil Nadu, while some others have taken advantage of the early flowering of the different dones in the orchard and collected seeds under open pollination clone-wise and raised seedlings for establishing progeny triaL One interesting observation reported by Kedharnath (1973) regarding some of the seed orchards pertains to early flowering observed in many clones, non-synchronous flowering among some ciones and production in general ofgood well-filled seeds with a very high percentage ofgermination. Intensive management of the orchards should be helpful in enhancing flower and seed production. Some of the states like Andhra Pradesh, Gujarat, Kerala, Madhya Pradesh, Maharashtra and Tamil Nadu have made good progress in the selection of plus trees and in establishing clonal seed orchards. Now efforts ate in progress to manage the orchards intensively so as to enhance flowe¡ and fruit production. Some of the teak plus trees ofTamil Nadu have been used as experimental material for gel electrophoresis studies to identify easterase bands (Kumaravelu 1979). Two other species, Tectona hamiltoniana Wall. and T. philippinensis Benth. and Horn. f. ex. Merr. are known under the genus Tectona. They are not very valuable as timber species. T. hamiltoniana has 36 as its somatic chromosome number. This species has been introduced in Dehradun from Burma. Trees of this species appear to be comparatively free from attacks ofleafskeletoniser and defoliator. Exploratory crosses were therefore attempted between T. grandis and T. hamiltoniana. This cross, however, yielded only shriveUed seeds and. failed to germinate. Embryological studies revealed Forest tree improvement in India 405 that fertilisation does take place in this cross but the hybrid embryo aborted very early. It should be possible to realisr this hybrid by •mploying embryo-culture technique. Grafting work carrir out with tbesr two species viz T. grandis as stock and T. hamiltoniana as scion showed that the grafts arr able to survive for about six years under Dehradun conditions and tben graft incompatibility manifests itself. This is not surprising, considering the fact that T. grandis has ring porus wood while T. hamiltoniana has diffusr porus wood. The same graft combination has been carried out also at K•mla Forest Research Institute, Peechi which ate 4 years old now and have grown well. It remains to be seen as to how soon late graft incompatibility will manifest itself in these. Ir is interesting in this connr to point out that studies carried by Gottwald and Parameswaran (1980) show that the general propr and anatomical features of the wood and bark, together with the leal trichomes, ate markedly different between T. grandis (of sr Tectona) on the one hand and T. hamiltoniana and T. philippinensis (of sect. Leiocarpae) on the other. Perhaps a taxonomical revision of the genus may suggest retention of a single species under the genus Tectona, viz T. grandis and the other two taxa T. philippinensis and T. hamiltoniana be shifted to another new genus or put under some other already r genus like for instancr Gmelina.

3.2 Eucalypts Three species of eucalypts in particular, Eucalyptus tereticornis, E. grandis and E. globulus ate in use for raising large scale plantations. E. globulus is mostly used in the Nilgiris in South India and no work on the genr improvement ofthis species has br initiated so far. In respr of the other two species a lot of research work has br162done for the genetic upgrading of the spr The provenancr of E. tereticornis usually referred to as,'Mysore Gum' or sometimes as 'Mysore hybrid' is the one that is in use in most of the states for raising large scale industrial plantations. It is also in use in . E. grandis is usually raised in higher ranges of west•rn ghats particularly in Kerala and to a small r in Tamil Nadu. These two spr191 ate worked on a short rotation of 8 yr. While in many areas ir has given good yields, in some areas, however, the yields have been rather poor. Work on provenancr testing is in progress in a number of states. Differences in tate of growth and suscr to Cylindrocladium blight has br reported by Jayashree et al (1984) from a study of 39 provenances rr162 15 species. A toxin bioassay method for assessing relative suscr of eucalypts to pink disease causr by Corticium salmonicolor has been reported by Sharma et al (1984). They screened 23 eucalypt entries and observed significant variation in their susceptibility. Plus tree selection and r of progeny t¡ has been taken up in E. grandis and E. tereticornis. Vegetative propaga tion by rooting of stem cuttings has not been very encouraging for large scale use. Clonal propagation by grafting has also not br very encouraging becausr of late manifestation of graft incompatibility. Thus, there has not been much enthusiasm for establishing clonal seed orchards. However, it is hoped that with the recent reports on the success achieved in obtaining plantlets from meristem culture in E. citriodora by Gupta et al (198 t) and in E. grandis by Lakshmi Sita et al (1984) there will be enthusiasm to use this approach for establishing clonal sr orchards. Valuable information on various genetic paramr has been reported from 406 S Kedharnath

E. tereticornis and E. grandis by Kedharnath and Vakshasya (1977), Kedharnath (1982a) and Krishnaswamy et al (1984). A number of spontaneously occurring interspecific hybrids have been identified and studied in India (Kedharnath 1980). These include E. camaldulensis x E. tereticornis, E. citriodora x E. torreliana, E. grandis x E. tereticornis. These hybrids manifest good tiybrid vigour for growth and volume production. It would be very beneficial to multiply them clonally and establish plantations using the tissue culture approach. Also, experimentally synthesised hyb¡ have been evaluated by Venkatesh and Sharma (1977). F 1 hybrids from some of the cross combinations exhibit good hybrid vigour.

3.3 Pines Till ver), re~ently only four spccies of pines were known in India--Pinus roxburghii Sarg., P. wallichiana Jack., P. kesiya Royle ex Gordon and P. gerardiana Wall. Now one more spr P. bhutanica Grierson, Long and Page, has becn recorded from Arunachal Pradesh. This spr was collected from Arunachal Pradesh by Naithani and Sahni in 1977 (Naithani and Bahadur 1981). The same species has been collected from Bhutan by Grierson et al (1980) and given the name P. bhutanica. It is a five-needle pino. Pinus roxburghii the low level pine confined to the monsoon belt of the outer Himalaya from Bhutan to North Eastern part ofWest Pakistan=i~ a valuable pine for its oleoresin and also timber. It grows in lower elevations gencrally up to 1830 m. Troup (1921) recognised nine provenances on the basis of growth characteristics. Considerable variation in oleoresin yield was found among the trees in the different provenances growing at New Forest, Dehradun in a provenance trial. In some provenances there were high-yielders of 4 to 7 kg. A programme of breeding for improving oleoresin yield was suggested by Kedharnath (1971). A number of plus trees spr for high resin yielding character were selected in the State of Uttar Pradesh. Clonal propagation tcchniques by Cleft grafting in the succulent region with 85-90 % graft take was worked out for this spr (Kedharnath et al 1979). Additionally, the grafted plants can be used for furtber clonal multiplication by air-layering. A very high pcrcentage of rooting response was obtained in airlayering trials carried out by Kedharnath and Dhaundiyal (1963). Successful rooting of stem cuttings using hormones and mist tent has also been reported from Himachal Pradesh Forest Department by Gupta (1979) and from the Forest Research Institute, Dchradurt; by Bhatnagar (1979). Plus trees in this spr have been selected based on characters such as good growth, stem form, straight cylindrical bole etc. by Khosla et al (1979) and Uniyal and Thapliyal (1979)~ The stage is now set for establishing clonal seed orchards. Additionally, valuable genetic information in this species has been reported by Snehalata Chawla (1977). Using ah open-pollinated progeny trial she assessed the natural variation in morphological, growth and wood characters. She has also obtained heritability estimates for the various traits and correlations both phenotypic and genetic amongst various wood characters. Studies on sensitivity of seeds of different seed origins of this species to acute gamma radiation have been reported by Upadhaya and Kedharnath (1974). When air dry seeds were used as experimental material the LP 50 for germination ranged between 3.31 and 9.12 KR. Two provenances were studied at 10 and 30% moisture content of sr In Forest tree improvement in India 407

one case the LD 50 came down to 7.50 KR from 9-12 KR when the moisture per cent of seeds was increased to 30 %. In the second case, the increase in moisture content to 30 % did not alter the LD 50. The karyotypes of some of the pine species of India have been examined in detail by Mehra and Khoshoo (1956), Kumar et al (1966) and Upadhaya and Kedharnath (1970). Pinus kesiya, the pine which occurs in the Khasi hiUs of Assam has also been taken up for genetic improvement at the Forest Research Institute, Dehradun. A pro venance experiment has been laid out at New Forest, Dehradun and is being assessed regularly. The same test had also been laid out in a number of states. Additionally,a cross between P. kesiya and P. merkussi was attempted over three years. Al1 the seeds obtained were sh¡ and they failed to germinate. In one year three viable seeds were obtained. But soon after germination the seedlings died. Thus there was no opportunity to confirm the hybridity of the seedlings. The blue pine of Himalaya, Pinus wallichiana, is a soft pine which is valued very much both for its timber and oleoresin. As a prelude to initiating genetic improvement work on this species variation has been studied by Dogra (1972). He has recognised seven altitudinal provenance types. Four of these are adapted to the outer moist and inner dry north-west Himalaya; and three to the outer wet, middle moist and inner dry eastern Himalaya. The major blue pine forests grow in Kashmir, Himachal Pradesh, Uttar Pradesh and Nepal. Bhutan is the major blue pine arca of the east. According to Dogra (1972) a weak reproductive barrier exists between the blue pine populations growing at lower and higher altitudes of both moist and dry zones but a strong reproductive barrier is functional between the moist and the dry arid zone blue pine of Himachal Pradesh. The additional imP0rtance of blue pine is its resistance to blister rust caused by Cronartium ribicola to which the two American pines Pinus strobus and P. monticola are highly susceptible. P. wallichiana has been used in crossing programme with the two American species cited above and resistant hybrids manifesting hybrid vigour have been realised in USA.

3.4 Introduced tropical pines Pinus patula, has been successfully introduced in West Bengal and at Kodaikanal and Ootacamund in Tamil Nadu. It is a promising species. But no work on the genetic upgrading of the spedes has been initiated so far in India. Pinus caribaea has been successfully introduced in a few States. The three varieties P. caribaea var. caribaea, P. caribaea var. hondurensis and P. caribaea var. bahamensis are included in the various trials. A number of provenances of these varieties are also under trial in a number of states. Field-grafting trials with P. caribaea has been carried out in India by Kapoor and Kedharnath (1976) and the time of the year best suited for field grafting has been ascertained. This should facilitate taking up work on the establishment of clonal seed orchard for this species.

3.5 Semul (Bombex ceiba L.) Semul is one of the valuable indigenous soft wood species which is in great demand for use in the match indust~y. The annual requirement of this wood by the match industry is of the order of 2 lakh tonnes. This tree belongs to the family Bombacaceae and has a 408 S Kedharnath

somatic chromosome number of ca.72. Semul is widely distributed on the Indian main land while the related species B. insigne Wall. is confined in its distribution to the Andamans, Western Ghats and Assam. The objective oftree improvement work in this species is to evolve va¡ which will be fast growing with good stem forro, narrow crown and without buttresses. In nature, in some arcas the trees are subject to heavy attacks by shoot borer fTonica niviverana Walk). So, incorporating resistance to this pest also forms one of the breeding objectives. Also, it is known that Semul from some arcas particularly that growing in Assam is valued more by the match industry because of the quality of wood. 'Plus trees' of Semul have been selected from those growing in Assam and in Uttar Pradesh and search for plus trees from other arcas are in progress. A small clonal seed orchard.has been established at Ranipur, 60 km from Dehra Dun using the simple grafting technique worked out for use with this species by Kedharnath and Venkatesh (1963). The grafts in the species flower the very next s~son after grafting if the scions had been carefully selected (Venkatesh and Arya 1967). Since each fruit contains 200-300 seeds and the percentage of germination of seed is very high, it has been › that half a hectare of seed orchard can yield enough seeds to plant up 500 hectares (Venkatesh 1970). The detailed observations taken on the flowering and fruiting in the different ciones in a half acre clonal seed orchard established at Ranipur, in Uttar Pradesh appear very promising from the point of view of good seed yield (Venkatesh and Arya 1967). The chromosome number in semul has been reported from meiotic and/or somatic counts by Baker and Baker (1968), Mehra and Sareen (1973) and Sareen et al (1980). Somatic numbers of 72, 92, and 96 have been recorded. This would mean that both hexaploids and octoploids are present in the species if we assume that the basic chromosome number is 12, It would be interesting to raise a progeny trial as well as a clonal trial from these different chromosome number trees and assess their perform- ance for growth and any other special attributes it may have such as resistance to drought, insect pests and also wood quality.

3.6 Red Sanders (Pterocarpus santalinus L.f.) Red Sanders is a very slow growing species con¡ to a small region in South India. It belongs to the family Papilionaceae and has 24 as its somatic chromosome number. The heavy, dark claret red heartwood has been in use for centuries for carvings, doll making etc. In recent years, a variant in this species which has wavy grained wood has leapt into sudden prominence because it is highly valued in the export market. Trees with this variant character occur at very low frequency in nature and tbey seem to show no apparent morphological differences by which they could be easily recognised from the normal grained trees. However, such individuals can be recognised from surrounding normal trees by blazing the sap wood, because the sap wood also shows the characteristic wavy grain. Since both normal and wavy grained trees oecur in the same general areas of dry sites with poor shallow soils, it is unlikely that this character is entirely controlled by environmental factors. Ifit is ah inhe¡ character, then the low frequency of its occurrence in nature would appear to indicate that the gene for this character is present in a low frequency in the population or the character is conditioned by multiple genes. Asa first step to increase the frequency of occurrence of trees with this variant trait, such trees have been identi¡ and assembled as grafts in a clone bank. These can then be asexually multiplied and a plantation establisbed. Seeds have been Forest tree improvement in India 409

collected from individual trees showing the wavy grain trait in the wood to raise half-sib progenies and sco¡ variation among and within the progenies in growth and other characters such as internode number and average internode length. It is anticipated that there will be segregation for two kinds of seedlings in each progeny---one normal looking and the other showing stunted growth with shorter internodes. This second category of seedlings may have a high probability of yielding trees with wavy grained wood. From the work carried out in Sweden on wavy grained trait in Birch, there is evidence that this trait is genetically eontrolled and the frequency of recovery of such plants in the progenies of trees with this trait varies. The variation of this trait from pith to periphery in a tree and between trees has been studied from wood core samples taken from trees at breast height. This revealed significant variation in the intensity of waviness from pith to periphery in individuals and also between trees. This information is now being used to select the most desirable trees for use in a seed orchard programme. It is anticipated that this orchard will produce seeds which would in turn yield plants that have a high probability of showing wavy grained wood. Variation in ¡ morphology and the growth of grafts from different dones have been studied (Kedharnath and Rawat 1976; Kedharnath et al 1976).

3.7 Poplars: (Populus spp.) Poplars, particularly dones of Populus deltoides, have a good future as a plantation crop in certain regions of North India. The ciones so far tried are those that have been tested and selected for site and climatic conditions obtaining abroad. Testing of some of these exotic ciones in the hope of identifying some amongst them as suitable for us is certainly a useful short cut approach to get something without much investment. But a more logical and realistic approach would be to develop our own ciones of the promising exotic species P. deltoides. An approach currently being taken up in Uttar Pradesh envisages (i) collection of seed resulting from open pollination on some of the good female ciones, (ii) attempt at controlled hybridisation between selected female and male ciones should they exhibit synchronised flowering. It is proposed to raise seedlings from the seeds resulting from the above two approaches and test them in the appropriate region and then the promising plants from amongst these will be cloned. Tests for resistance to important diseases and in~ect pests will also be carried out. While this work with exotic ciones ofP. deltoides progresses, it has also been planned to work on the genetic upgrading ofP. ciliata and P. gamblei, two ofour native poplars. A programme of genetic improvement work has been proposed for the poplars in India by Kedharnath (1979). Khurana and Khosla (1982) have been active in the selection of desirable phenotypes in P. ciliata and studying their variation in provenance testing. Khosla et al (1979) have also assessed the sex ratio in natural population of this species and studied the correlation between the sex of the tree and its growth.

4. Strategies for the future

Problems of immediate importance and finding solutions to them certainly deserve high priority and in this context tree breeding programmes had set high priority for the selection of plus trees and assembling them in clonal seed orchards so that as soon as the 410 S Kedharnath orchards started producing regular crops of seeds, genetically improved planting mate¡ become available for raising the new plantations. That would mean immediate gains. But it is also necessary to think of the longterm goal and plan for building material for advance generation or multigeneration breeding programmes. In de- veloped r241 where forest tree breeding programmes have been in operation for a long time tree breeders have given much thought in this direr For example, Bourdon et al (1977) examined a wide range of alternative mating designs for va¡ purposes including estimates of variances and combining abilities, development of breed populations and production of seed. They found that no single design was best for all purposes and no single purpose will be served by only one design. Ar to Lindgren (1977) reasonably good progeny tests can be made with a limited number of trees in any of the several designs including common testers, partial diallels, polyr and pollinations in seed orchards. Strategies suggested for the development of long-term genetic improvement programmes by different experts in the field of tree breeding differ quite markedly. However, they all agree on the need to separate the short-term function of seed produr from the long-term goal of developing and maintaining broad-based genetic populations for future advances in the tree improvement. The commonest example of a production population is a seed orchard. In the orchard we generally tend to increase the genetic gain by inr the selection differential. This may appear asa conflict to maintaining a broader genetic base for future breeding work. In the past there has been rigorous seler of plus trees so that only the best or more outstanding individuals were included in the seed orchard. However, the present tendency is to select a large number of good trees (rather than a few super trees) in the first round of seler of plus trees. Thus itis exper that it would not only ensure a broad genetic base than before but would also facilitate a reasonable level of improvement in the second and subsequent generations (Pederick and Griffin 1977). It is very satisfying to know that today amongst foresters there is an inr appreciation of the role and potentiality of genetics and tree breeding in maximising production from the forest plantations.

References

Baker H G and Baker J 1968 Chromosome numbers in Bombacaceae; Bot. Gaz. 129 294-296 Bhatnagar H P 1979 Rooting of cuttings of chirpine (Pinus roxburohª personal communication Bourdon R D, Shelbourne C J A and Wilcox M D 1977 Advanced selecfion strategies; Proc. 3rd World consultation on Forest Tree Breeding Canberra, Australia 2 1133-1148 Dogra P D 1972 Intrinsic qualities, growth and adaptation potential of Pinus wallichiana; U S Dept. Agric. For. Serv. Misc. Pub. No. 1221 163-178 Gottwald H and Parameswaran N 1980 Anatomy of wood and bark ofTectona (verbenaceae) in relation to taxonomy;, Bot. 3ahrb. Syst. 101 363-384 Grierson A J C, Long D G and Page C N 1980 Notes relating to the flora of Bhutan: (III) Pinus bhutanica; a new 5-needle pine from Bhutan and India; Notes R. Bot. Gard. Edinburgh 38 297-310 Gupta M P 1979 Rooting of cuttings in Chirpine (Pinus roxburohiO personal communication Gupta P K, Mascarenhas A F and Jagannathan V 1981 Tissue culture of ForestTrees: Clonal propagation of matª trees of Eucalyptus citriodora Hook., by tissue culture; Plant Sci. Lett. 20 195-201 Gupta P K, Nadgir A L, Mascarenhas A F and Jagannathan V 1980 Tissue culture of Forest Trees: Clonal multiplication of Tectona grandis L. f. (teak) by tissue culture; Plant Sci. Lett. 17 259-268 Jayashree M C, Madhavan Nair J, Arvind D D and Ramaswamy V 1984 Relative susceptibility ofeucalypts provenances to Cylindrocladium blioht. Paper presented at the National Seminar on eucalypts held at Peechi. Kerala Forest tree improvement in India 411

Kapoor M L and Kedharnath S 1976 Field grafting t¡ with Caribaean pine; lndian For. 102 279-82 Kedharnath S 1971 Evolving high Oleo-resin yielding strains of chirpine (Pinus roxburohii) through breeding; Proc. Seminar, Sympine, Delhi, India DI-D5 Kedharnath S 1973 Forest tree breeding in India; Proc. First Forestry Conf., Dehradun, India Kedharnath S 1974 Genetic improvement of some forest tree species in India; lndian J. Genet. Plant Breed. A34 367-374 Kedharnath S 1979 Evolving geneticaUy improved clones of poplars for planting in India; Proc. Symp. on , manaoement utilisation of poplars Srinagar 150-161 Kedharnath S 1980 Genetic improvement offorest trees, position paper presented at the Second Forestry Conference, Dr India Kedharnath S 1982 Plus tree selectiotr--a tool in forest tree improvement in lmprovement offorest (ed.) P K Khosla p 13-20 Kedharnath S 1982a Crenetic variation and heritability of juvenile height growth in Eucalyptus grandis; 3. Tree Sci. 1 46-49 Kedharnath S, Chac,ko V J, Gupta S K and Matthews J D 1963 Geographic and individual tree variation in some wood characters of teak (l'ectona grandis L.f.) Fibre length; Silvae Genet. 12 181-186 Kedharnath S and Dhaundiyal R P 1963 Preliminary observations on air-layering in Pinus roxburohii and P. caribaea lndian For. 89 219-221 Kedharnath S, Kapoor M L and Vakshasya R 1979 A note on field grafting in chirpine; lndian For. 105 301-304 Kedharnath S and Mathews J D 1962 Improvement of teak by selection and breeding; lndian For. 88 277-284 Kedharnath S and Pratap Singh 1975 Studies on natural va¡ in susceptibility of Tectona to leaf skeletoniser, Pyrausta machaeralis paper presented at FAO/IUFRO Symp. on Forest diseases and insects; New Delhi--India Kedharnath S and Raizada M B 1961 Genetics and Forest tree breeding; Proc. lOth Silvicultural Conf Dehradun, India 203-214 Kedharnath S and Rawat M S 1976 Studies on va¡ in fibre morphology in wavy-grained and straight- grained trees of Red Sanders; lndian For. 102 441-446 Kedharnath S, Rawat M S, Uniyal D P and Lakshmikantham D 1976 Studies on field grafting and the growth of the grafts in Red Sanders; lndian For. 102 761-765 Kedharnath S, Rawat M S and Chauhan V S 1970 Early growth performance of twenty ciones of tr (Tectona grandis L.f.) in a seed Orchard, Proc. Seminar-cum-workshop on Genetic improvement offorest tree seeds in India, Dehradun 86-89 Kedharnath S and Vakshasya R 1977 Estimate of components of variance, heritability and correiations of some growth parameters in Ecalyptus tereticornis; Proc. 3rd World Consult. Forest tree Breeding, Canberra, Australia 2 667-676 Kedharnath S and Venkatesh C S 1963 Grafting as an aid in the breeding of teak (l'ectona grandis L.f.) and Semul (Salmalia malabarica Schoot et Endl.); Proc. World consultation on forest genetics and tree improvement, Stockholm, Sweden, 2 5/6-12 Khosla P K, Dhall S P and Khurana D K 1979 Studies in Populus ciliata Wall. Ex. Royle. I. Correlation of phenotypic observation with sex of trees; Silvae Genet. 28 21-23 Khosla P K, Seghal R N and Sagwal S S 1982 Seed coilection and seed certification with referente to Pinus roxburghii Sarg. in Himachal Pradesh, in lmprovement offorest biomass (ed.) P K Khosla, Pub. Indian Soc. ~of Tree Scientists Khurana D K and Khosia P K 1982 Concept of provenance testing and provenancr t¡ in Populus ciliata Wall. Ex. Royle. in lmprovement offorest biomass (cid.) P K Khosla, Pub. Indian Soc. Tree Scientists. Krishnaswamy S, Vinaya Rai R S and Srinivasan V M 1984 Studivs on variante components and heritabifity in one-parent families of Eucalyptus tereticornis. Paper presented in the Nationai Sr on Eucalypts held at KFRI Peechi, Kerala Kumar S, Bansal H C, Singh D and Natarajan A T 1966 Consistency of karyotypes and ciassification and identification of chromosomes of genus Pinus; lndian J. Genet. 26 311-316 Kumaravelu G 1979 Clonai identification ofTectona grandis by Isoenzyme studies; lndian For. 105 716-719 Lakshmi Sita G, Sobha Rard and Sankar Rao K 1984 Propagation of Eucalyptus grandis by tissue c ulutre. Paper presented in the National Sr on Eucalyptus held at KFRI Peec¡ Kerala Lindgren D 1977 Genetic gain by progeny testing asa function of mating design and cost, Proc. 3rd World consultation on Forest Tree Breedin#, Canbr Austral/* 2 1223-1235 Mehra P N and Khoshoo T N 1956 Cytology of Conifvrs; lndian J. Genet. 54 165--180 Mehra P N and Sareen T S 1973 Cytoiogy of some Himalayan trer Thalamiflorae; Silvae Genet. 22 66-70 412 S Kedharnath

Naithani H B and Bahadur K N 1981 Observations on extended distribufion ofnew and rare taxa of north- eastern India with sper reference to Arunachal Pradesh; lndian For. 107 712-724 Pede¡ L A and Griffin A R 1977 The genetic improvement of radiata pine in Australasia, Proc. 3rdWorld Consultation on Forest Tree Breedino, Canberra, Australi& 2 561-572 Rawat M 5 and Kedharnath S 1968 Field grafting and budding in teak (Fectona orandis L.f.) lndian For. 94 260-261 Sareen T 5, Uppal (Mrs) 5 and Kant 5 1980 Chromosome numbers of some woody angiosperms; lndian 3. For. 3 73-77 Sharma J K, MarŸ Florence E J, Sankaran K V and Mohanan C 1984 Toxin bioassay--a rap¨ methodfor assessino relative susceptibility of eucalypts ooainst pink disease, Paper presented in the National Seminar on Eucalypts held at KFRI Peechi, Kerala Snehalatha Chawla 1977 Studies on variatian, inheritance and correlation amongst growth morphological and wood characters in chirpine ( Pinus roxburohit) Ph.D thesis, Garhwal University Troup R S 1921 The silviculture oflndian trees 3 1013-1095 Uniyal D P and Thapfiyal R C 1979 Selec• of plus trees ofchirpine (Pinus roxburohii ) in the Tongs division of Uttar Pradesh (personal communication) Upadhaya L P and Kedharnath S 1970 Karyotype offour species ofpines naturally occurring in India; lndian For. 96 657-667 Upadhaya L P and Kedharnath S 1974 Sensi ofseeds ofchirpine ofdifferent seed origin to acute gamma irradiation; lndian J. Genet. Plant Breed. A34 393-399 Venkatesh C S 1970 Genetic quality control of Semui for the Indian Match ; Van Figyan. 8 93-95 Venkatesh C S and Arya R S 1967 Observafions on the flowering and fruiting behaviour of Semul gratis; Ind/an For. 93 586-587 Venkatesh C S and Sharma V K 1977 Differential heterosis in reciprocal interspecific crosses of Eucalyptus camaldulensis and E. tereticornis, Proc. 3rd World Consultation of Forest Tree Breedino, Canberra, Australia, 2 677-682 Levels of Genetic Variation in Trees: Influence of life history characteristics1

J. L. Hamrick,2 J. B. Mitton,3 and Y.B. Linhart3

Abstract: In a previous study, levels of genetic variation, as measured by isozyme analyses, were compared for 113 taxa of vascular plants. Each species was classified for 12 life history and ecological traits and three measures of genetic variation were calculated. Plants with large ranges, high fecundities, an outcrossing mode of reproduction, wind pollination, a long generation time, and from habitats representing later stages of succession tended to have more isozyme variation than species with other combinations of characteristics. This paper discusses the results of the previous study and examines the available isozyme data for similar trends in forest trees. Special consideration was given to differences in genetic variation among 20 species that have many of their life history characteristics in common. Successional stage, habitat type, cone type and historical events were associated with differences in genetic variation among the conifer species. These results are discussed in terms of expectations from current population genetics theory.

orest trees have been the subject of many Hamrick 1979, Hamrick and others 1979). Generally, these F quantitative genetic investigations (Libby and others reviews have concluded that plants contain somewhat 1969, Stern and Roche 1974). These studies have more variation than invertebrate animals and considerably concentrated on morphometric and physiological charac­ more variation than most vertebrates. Furthermore, teristics such as survival, growth initiation, height and different plant species contain varying amounts of genetic diameter growth, hardiness to environmental stress, and variation. Trees, for example, have been found to contain various leaf, stem, fruit, and wood characteristics. As a significantly more variation than herbaceous plants result, the distribution of quantitative genetic variation is (Hamrick 1979, Hamrick and others 1979). better understood in certain tree species than in most other This paper reviews the results of a previous study naturally occurring plants. (Hamrick and others 1979), which compared the isozyme Studies that use biochemical techniques to measure variation of plant species with different combinations of genetic variation in forest trees generally have lagged life history traits and examines the available isozyme data behind those that use quantitative traits. A few workers for similar trends in forest trees. Specifically, we address have used secondary plant compounds, such as terpenes, in the following questions: studies of species (Zavarin and Snajberk 1969, Zavarin and • Do plant species with certain combinations of life others 1969), racial (Smith and others 1969) and history traits contain higher levels of genetic variation? population (Adams 1975a, 1975b) differentiation. But, • Does being a woody plant directly affect levels of because the genetic basis of variation in these compounds is isozyme variation? poorly understood, they are of limited use in population • Can differences in genetic variation among tree species genetic studies. Since the early 1970's, electrophoretic be explained by their life history and ecological techniques have been used in genetic studies of forest tree characteristics? populations. These techniques offer a number of • advantages over other biochemical or quantitative PROCEDURES approaches: (a) genetic inheritance of electrophoreticaIly­ detectable traits can be easily demonstrated; (b) most In a previous study (Hamrick and others 1979), we isozyme loci are codominant and gene frequencies can be examined the relationship between 12 life history and calculated without the necessity of genetic crosses; (c) ecological traits and the levels of genetic variation estimates of genetic variation can be compared directly maintained within populations of 113 taxa of plants. Each between populations or between species. The relatively few species was classified for taxonomic status, geographic isozyme studies of plant populations have been the subject range, generation length, mode of reproduction, mating of a number of recent reviews (Gottlieb 1977, Brown 1979, system, pollination mechanism, fecundity, seed dispersal mechanism, chromosome number, successional stage, habitat type, and cultivation status. Where the data 1 Presented at the Symposium on Isozymes of North American Forest allowed, three measures of intrapopulation genetic Trees and Forest Insects, July 27, 1979, Berkeley, Calif. 2Associate Professor of Biology, University of Kansas, Lawrence, variation were calculated for each species: the percent of Kans. polymorphic loci per population (P), the mean number of 3 Associate Professor of Biology, University of Colorado, Boulder, alleles per locus (A), and a polymorphic index (PI). The PI Colo.

35 Table 1—Levels of variability among categories of 12 life history and ecological traits. Weighted means and standard errors are given for each measure of variability. Differences in PI between categories are tested by ANOVA. Statistical significance levels are given in parentheses (from Hamrick and others 1979)

Polymorphic Alleles per Polymorphic Variable Species Loci loci (P) locus (A) Index (PI) x S.E. x S.E. x S.E. Percent

Taxonomic status (P <0.01) Gymnospermae 11 9.2 67.01 7.99 2.12 0.20 0.270 0.041 Dicotyledoneae 74 11.4 31.28 3.31 1.46 .06 .113 .014 Monocotyledoneae 28 11.6 39.70 6.02 2.11 .19 .165 .026 Geographic range (P < 0.05) Endemic 17 15.1 23.52 5.06 1.43 0.11 0.086 0.019 Narrow 22 11.4 36.73 6.01 1.60 .14 .158 .030 Regional 39 8.3 55.96 5.13 1.85 .10 .185 .025 Widespread 35 12.5 30.36 5.03 1.58 .15 .120 .021 Generation length (P <0.001) Annual 42 11.2 39.47 4.32 1.72 0.11 0.132 0.017 Biennial 13 17.2 15.78 5.12 1.26 .09 .060 .020 Short-lived 31 12.0 28.09 5.06 1.46 .09 .123 .023 perennial Long-lived 27 7.6 65.77 5.08 2.07 0.13 0.267 0.027 perennial Mode of reproduction N.S. Asexual 1 8.0 50.00 0.00 1.91 0.00 0.139 0.000 Sexual 95 11.7 35.64 3.03 1.63 .07 .135 .012 Both 17 8.9 41.71 8.12 1.67 .14 .185 .034 Mating system (P < 0.01) Primarily selfed 33 14.2 17.92 3.21 1.27 0.06 0.058 0.014 Mixed 42 8.6 14.16 4.89 1.76 .10 .181 .022 Primarily outcrossed 36 11.3 51.07 4.95 1.85 .12 .185 .022 Pollination mechanism (P <0.001) Selfed 33 14.2 18.99 3.51 1.31 0.07 0.058 0.028 Animal 55 9.5 38.83 3.94 1.55 .07 .130 .015 Wind 23 10.7 57.45 6.29 2.27 .17 .264 .028 Fecundity (P < 0.001) <102 21 12.0 40.06 6.45 1.72 0.16 0.127 0.026 102 to 103 27 12.4 26.35 3.46 1.44 .07 .096 .013 103 to 104 22 11.9 36.98 6.37 1.64 .10 .199 .034 >104 40 9.8 67.99 5.99 2.27 .17 .286 .033 Seed dispersal N.S. mechanism Large 27 11.4 37.42 3.73 1.76 0.156 0.0230 .16 Animal-attached 16 11.1 28.79 5.55 1.55 .08 .092 .020 Small 26 12.4 32.98 5.10 1.51 .09 .118 .018 Winged or plumose 21 12.2 44.91 7.27 1.86 .13 .188 .029 Animal ingested 20 7.0 32.98 8.25 1.43 .10 .132 .036 Chromosome number (P<0.01) 10 to 20 50 13.1 35.52 3.61 1.55 0.08 0.111 0.014 22 to 30 44 10.0 37.41 5.35 1.73 .10 .175 .023 >30 16 8.9 41.65 7.20 2.10 .14 .224 .030 Stage of succession (P <0.01) Weedy and early 54 12.5 29.67 3.82 1.60 0.08 0.116 0.015 Middle 49 9.7 37.90 4.43 1.56 .08 .137 .019 Late 10 12.0 62.76 5.28 2.14 .19 .271 .038 Habitat type N.S. Xeric 4 8.8 15.39 8.20 1.11 0.09 0.048 0.040 Submesic 19 10.5 43.68 4.86 1.66 .08 .140 .020 Mesic 82 11.4 36.01 3.61 1.65 .07 .146 .016 Hydric 8 13.0 27.71 10.33 1.59 .22 .145 .050 Cultivation status N.S. Cultivated 21 6.8 38.99 7.15 1.61 0.12 0.172 0.032 Noncultivated 89 12.6 36.10 3.12 1.63 .07 .136 .013 Both 3 2.7 50.00 — 1.75 .21 .209 .035

36 is equivalent to the Hardy-Weinberg heterozygosity Multivariate techniques were used to identify groups of (Hamrick and Allard 1972). variables varying in concert, and to assess the covariation For each category of the 12 life history variables of genetic variation with ecological and life history weighted (by the number of loci), means were calculated variables. Approximately one-half of the correlations for P, A, and PI, and mean PI values were compared for among the ecological, life history, and genetic variables heterogeneity by single classification ANOVA. Multivar­ were statistically significant. Pollination mechanism, iate statistics were used to determine correlations among mating system, and fecundity had the highest correlations traits and to test whether combinations-of life history with the genetic variables. Additional multivariate variables influence genetic variation. The data were analyses demonstrated that only the first two principal analyzed first with a principal components analysis to components contributed to our understanding of the genetic describe the major patterns of covariation. Associations variation among these species. The first principal noted in this analysis were explored further with a stepwise component, which had high loadings from the three genetic multiple regression. variables, generation length, mating system, pollination Differences in genetic variation among tree species was mechanism, fecundity, seed dispersal and successional the subject of a second analysis. This analysis included status explained 30 percent of the variation. The second studies not available earlier but was limited to studies that principal component, which had high loadings from the used a wide variety of enzyme systems. Values of P, A, and genetic variables, taxonomic status, seed dispersal PI were calculated for each species and weighted means mechanisms, and successional stage explained 16 percent were calculated for each category of six variables. of the variation. These results were consistent with the Multivariate analyses were not used since most of the tree univariate analyses and indicated that species with large species were conifers and, therefore, similar for many of ranges, high fecundities, an outcrossing mode of their life history traits. reproduction, wind pollination, a long generation time, and from habitats representing later stages of succession had more genetic variation than did species with other combinations of traits. It is worth noting that forest trees in RESULTS general and conifers in particular combine many of the characteristics that are associated with high levels of Isozyme Variation in Plants genetic variation. To determine which ecological and life history traits The mean values of P, A, and PI obtained by pooling the were most closely associated with genetic variation we 113 taxa in our original study were: P= 36.8 percent, A = employed a stepwise multiple regression analysis in which 1.69, and PI = 0.141. When compared with animals, plants PI was the dependent variable and the 12 ecological and life tend to have levels of genetic variation that are roughly history traits were independent variables. Only two equivalent to the invertebrates (P= 46.9 percent, PI= 0.135 variables were significant—pollination mechanism and [Selander 1976]; P = 39.7 percent, PI = 0.112 [Nevo 1978]) fecundity. Species that are wind pollinated and highly but are considerably higher than those of vertebrate species fecund were the most genetically variable. (P = 24.7 percent, PI = 0.061 [Selander 1976], P = 17.3 percent, PI = 0.036 [Nevo 1978]). Variation Among Tree Species

Genetic Variation and Life History Traits The separate analyses of the tree data produced some noteworthy results. First, the mean level of variation Statistically significant differences (P<0.05) were found within populations of coniferous trees was lower than that among categories of eight life history or ecological traits reported previously (Hamrick and others 1979). In the (table 1). Gymnosperm species tended to have more earlier review (Hamrick and others 1979), the inclusion of variation than angiosperms while regionally distributed studies based solely on polymorphic loci tended to increase species had more variation than those with other estimates of the mean levels of variation. Also, many of the geographic ranges. We had expected species with the earlier tree studies used enzyme systems such as esterases widest ranges to contain the most variation. Many species and peroxidases that are often highly variable. The data in in this category, however, are weedy or early successional the present analyses are relatively free of these biases and species that tend to have less variation. Long-lived should be more representative of the actual levels of genetic perennials and plants with mixed or primarily outcrossed variation in tree populations. It is significant, therefore, mating systems contained at least twice the variation of that the mean levels of variation in tree populations species in other categories. Within the outcrossed species, continue to exceed that of herbaceous species by more than wind-pollinated species had more variation than those with 60 percent. animal pollination. Species with high fecundities and high A second important result is the continued observation of chromosome numbers also had more variation as did interspecific differences in genetic variation. The PI varies species of the later stages of succession. from 0.000 for red pine (Pinus resinosa Ait.) to 0.364

37 Table 2—Summary of electrophoretic studies of genetic variation in trees. P = percent of polymorphic loci, A = alleles per locus, PI = polymorphic index

Species1 Location Loci Populations P A PI Source

A. Gymnosperms Abies balsamea New Hampshire 14 1 64.0 1.86 0.150 Neale (1978) Picea abies Sweden 12 4 91.7 3.54 .341 Lundkvist (1979) P. sitchensis Rangewide 24 10 — 1.90 .150 Yeh, F. (unpubl.) Pinus aristata Rangewide 22 5 46.4 1.55 .139 Hiebert, R.D. and Hamrick, J.L. (unpubl.) P. attenuata Rangewide 22 10 73.0 2.09 .140 Conkle, M.T. (unpubl.) P. balfouriana Rangewide 23 4 57.6 1.61 .208 Hiebert, R.D. and Hamrick, J.L. (Unpubl.) P. banksiana Michigan 21 1 28.6 — .083 Snyder, T. (unpubl.) P. contorta British Columbia 27 17 — 1.90 .150 Yeh, F. (unpubl.) P. contorta California 37 1 89.2 2.78 .190 Conkle, M.T. (unpubl.) P. contorta Colorado 26 1 42.0 1.42 .160 Hamrick, J.L. (unpubl.) P. lambertiana Rangewide 20 — 80.0 2.85 .260 Conkle, M.T. (unpubl.) P. longaeva Rangewide 14 5 78.6 2.35 .364 Hiebert (1977) P. jeffreyi California 44 1 — 2.90 .260 Conkle, M.T. (unpubl.) P. muricata Northern California 17 1 65.0 1.53 .090 Millar, C. (unpubl.) P. ponderosa Eastern Colorado 22 7 68.4 2.00 .226 Hamrick, J.L. (unpubl ) P. pungens Rangewide 15 3 40.0 1.33 .144 Feret (1974) P. resinosa Rangewide 9 5 0.0 1.00 .000 Fowler and Morris (1977) P. rigida Rangewide 21 11 78.8 2.19 .144 Guries, R. and Ledig, T. (unpubl.) P. strobus Seed Orchard, 17 — 52.9 2.06 .330 Eckert and others (1980) New Hampshire P. taeda North Carolina 11 1 100.0 3.73 .340 Conkle, M.T. (unpubl.) P. taeda Superior Trees 30 — 93.3 3.87 .260 Conkle, M.T. (unpubl.) P. taeda Seed Orchard, 15 — 80.0 2.93 .200 Adams and Joly2 South Carolina Pseudotsuga British Columbia, 21 11 — 2.23 .180 Yeh, F. (unpubl.) menziesii Interior P. menziesii Coastal 21 11 — 2.23 .150 Yeh, F. (unpubl.) P. menziesii California Coastal 11 9 74.2 3.17 .332 Morris, R. (unpubl.) P. menziesii California Interior 17 1 100.0 1.78 .330 Conkle, M.T. (unpubl.) P. menziesii Eastern Colorado 22 5 64.0 1.86 .264 Hamrick, J.L. (unpubl.) Sequoiadendron Rangewide 8 34 50.0 2.63 .155 Fins, L. (unpubl.) giganteum Mean 20.1 67.7 2.29 .207 ±4.9 ±0.14 ±.017

B. Angiosperms Persea americana 10 80.0 1.90 .195 Torres and others (1978) cultivars Phoenix dactylifera 7 100.0 2.00 .332 Torres, A.M. (unpubl.) cultivars Mean 8.5 88.23 1.94 .251 ±10.0 ±0.05 ±.028

1Only those studies that used a variety of enzyme systems are included. 2Adams, W.T., and R.J. Joly. Allozyme studies in loblolly pine seed orchards: clonal variation and frequency of progeny due to self-fertilization (Manuscript in preparation.) for Great Basin bristlecone pine (P. longaeva D.K. Bailey). genetic variation as conifers, final conclusions must await Since the 20 conifer species have many of their life history studies of naturally occurring angiosperm trees. characteristics in common, only six traits were available to The geographic range of each conifer was classified as explain the differences in genetic variation observed endemic, narrow, or regional (table 3). Most species had a among species—taxonomic status, geographic range, stage regional distribution with only foxtail pine (Pinus of succession, habitat type, cone type, and U.S. balfouriana Grev. & Balf.) and giant sequoia (Sequoia­ distribution. dendron giganteum [Lindl.] Buchholz) classified as The present data (table 2) are inadequate to determine endemics. The differences in PI between these categories whether differences between angiosperm and gymnosperm were small and were not significant (P < 0.50). trees were significant. The two studies of angiosperm trees The ability of a species to successfully reproduce in its that met our criteria involved commercial cultivars of fruit own shade and in the absence of environmental trees. Although angiosperm trees appear to have as much disturbance was used to classify the species into one of

38 Table 3—Levels of variability among categories of five life history and ecological variables for 28 conifer studies. Weighted means and standard errors are given. Differences in PI between categories are tested by ANO VA. Significance levels are indicated in parentheses

Mean Polymorphic Index Variable Species1 Studies loci (PI)

Mean S.E. Geographic range (P < 0.50) Endemic 2 2 15.5 0.194 0.052 Narrow 8 8 22.2 .197 .032 Regional 10 18 19.7 .207 .023 Stage of succession (P < 0.10) Early 7 10 19.9 0.161 0.030 Middle 7 11 19.8 .241 .022 Late 7 7 20.8 .204 .035 Habitat type (P < 0.10) Xeric 3 3 19.3 0.194 0.075 Submesic 11 15 21.7 .186 .021 Mesic 6 10 17.9 .238 .026 Cone type (P < 0.01) Closed cone 6 7 21.3 0.132 0.012 Open cone 15 21 20.4 .226 .020 United States distribution (P<0.30) Northeastern 5 5 16.4 0.152 0.054 Southern 2 4 17.8 .235 .042 Western 12 18 22.1 .204 .018

1 Some species may be represented in more than one category. three stages of succession (table 3). More variation (P < cones and a southern or western distribution have more 0.10) was observed in populations of middle and late genetic variation than species with alternate combinations successional species and less in populations of pioneering of characteristics. species. Only the presence of loblolly pine (P. taeda L.) in the early category prevented the results from being statistically significant. DISCUSSION Differences among the habitat categories approached statistical significance (P < 0.10), but no definite trend can A significant proportion of the differences in genetic be seen (table 3). Differences between the submesic and variation among plant species is accounted for by variation mesic categories suggest a pattern but with only two species in their life history and ecological characteristics. Species in the xeric category no definite conclusions can be drawn. with large ranges, high fecundities, an outcrossing mode of Studies of genetic variation in conifer species that are reproduction, wind pollination, a long generation time and adapted to drought conditions, such as the piñon pines and from habitats representing later stages of succession have junipers, should yield valuable information on this higher amounts of genetic variation. This result is generally question. consistent with that predicted by population genetics Species with closed cones or ecotypes of species that theory. Plants, in general, might be expected to maintain have closed cones were grouped together. These species had high levels of genetic variation within their populations significantly less variation (P<0.01) than open-coned since their sessile nature often leads to the evolution of species (table 3). locally adapted ecotypes (Antonovics 1971, Bradshaw Species which inhabit different geographic regions 1972, Jain and Bradshaw 1966). Also, long-lived plant within the United States may also have different levels of species with large ranges and high fecundities typically genetic variation. The northeastern species (Abies have large, stable populations. Such populations are balsamea [L.] Mill., Pinus banksiana Lamb., P. resinosa resistant to chance fluctuations in gene or genotype Ait., P. rigida Mill., and P. strobus L.) have somewhat less frequencies and should maintain more variation than variation (P < 0.30) while the southern (Pinus taeda L. and populations that experience large fluctuations in size. P. pugens Lamb.) and the western species have higher Longevity also ensures the representation of many cohorts values. within a population. If different alleles or genotypes are To summarize the results of the analyses on trees, species favored during the establishment phase of each cohort, of later successional stages, mesic habitat types, with open individuals that survive to maturity will maintain a genetic

39 "record" of these evolutionary events. Their continued Buchholz, are long-lived and could maintain genetic survival would retard the loss of genetic variation. Greater variation by this mechanism. longevity could be especially effective in maintaining Trends observed for stage of succession, habitat type, genetic variation in later stages of succession because and cone type in the conifer data are generally consistent individuals are continuously becoming established in a with previous results (Hamrick and others 1979); conifers highly complex biotic environment. It should be noted, from mid- and late-successional stages, mesic habitats, and however, that the seed carryover abilities of many annuals with open cones tend to have higher levels of genetic and short-lived perennials could produce a similar genetic variation. Trees of early successional stages must adapt to record. Finally, high rates of fecundity, outcrossing and relatively homogeneous environmental conditions that are wind pollination ensure large neighborhood sizes and the influenced primarily by physical factors. Coupled with production of a variety of genotypes through recombina­ their colonizing habit, such uniform selection pressures tion. Natural selection could act to maintain this variation might lead to a reduction in genetic variation. As through the evolution of locally adapted ecotypes or succession proceeds, the biotic environment becomes through various types of . increasingly important and, as a result, habitats become The existence of high levels of genetic variation in forest more complex and heterogeneous. Such complex tree populations can be explained primarily by their life environments may select for the maintenance of higher history and ecological characteristics. The question levels of genetic variation. A similar explanation may remains, however, of whether woodiness has a direct effect apply to the higher levels of variation observed in the more on the levels of genetic variation that are maintained. mesic adapted trees. Current evidence, although scanty, indicates that herba­ The lower levels of variation found in the closed-cone ceous species with life history or ecological traits similar to pines could result from a combination of factors. First, the those of trees also have high levels of genetic variation. closed-cone species are adapted to habitats which This would argue against woodiness itself as affecting the experience periodic catastrophic and are, maintenance of genetic variation. An alternate argument therefore, almost always members of early successional can be made on the basis of differences in life forms communities. Such species germinate and grow under (Raunkiaer 1934) between woody and herbaceous plants relatively uniform environmental conditions. Also, the (Hamrick 1979). Woody plants are phanerophytes and closed cone habit and short life expectancy ensures even- maintain their apical meristems above ground. Their apical meristems, therefore, are exposed to environmental aged stands. The genetic record of the long-lived, uneven fluctuations throughout the year. Herbaceous perennials aged species is not, therefore, an important factor in the are either chamaephytes, hemicryptophytes or crypto­ maintenance of genetic variation in closed-cone species. phytes whose apical meristems are located at or below the The geographic region to which conifer species are soil surface during periods of severe environmental stress. native also seems to affect genetic variation. Four of the The moderating effects of snow, soil, or litter cover may, in five northeastern conifers maintain less variation than their essence, reduce selection pressures for the internal southern and western counterparts. Two factors may help buffering thought to be produced by increased heterozy­ to explain this observation. First, it could be argued that gosity (Lerner 1954). If this argument is valid we would western habitats are environmentally more heterogeneous expect to find differences in genetic variation between than those in the northeast. Support for this argument populations of herbaceous and woody plants that occur in comes from the greater diversity of community types in the the same habitats and have similar life history traits. Also, West (Küchler 1964). The high variation in the two we would expect to find an increase in heterozygosity in southern species contradicts this argument, however. A older age classes and in temporally fluctuating environ­ second factor to consider is the evolutionary history of ments. An increase in heterozygosity in older age classes each species. The northeastern species have been exposed was demonstrated in a population of Liatris cylindracea to numerous continental glacial events. Glaciation may Michx. (Schaal and Levin 1976). A further test of this have greatly reduced the ranges of the northeastern species argument would be provided by surveying a wider variety and reduced population sizes and genetic variation. of woody trees and shrubs. Although the western species have also been exposed to Differences in genetic variation among the conifer changes in climatic factors and shifts in their ranges, the species surveyed are somewhat more difficult to explain. more varied topography of the West may have provided a This is because the 20 conifer species have many of their life greater variety of refugia. In fact, some of the western history characteristics in common. Although only one of species expanded their ranges during glacial periods (for the six traits — cone type — was statistically significant, example, Pinus longaeva D. K. Bailey4). geographic range was the only trait that failed to have large In conclusion, a thorough knowledge of a species' life differences among the mean PI values. This result is not history and ecological characteristics is essential if its surprising since endemic and narrowly distributed tree genetic structure is to be understood. Much of the species often have rather large population sizes where they heterogeneity in genetic variation seen among species can occur. Also both of the endemic species, Pinus balfouriana Grew. & Balf. and Sequoiadendron giganteum (Lindl.) 4Personal communication from P.V. Wells,

40 be explained by such considerations. A significant amount of unexplained heterogeneity remains, however. Past Hamrick, J.L., and R.W. Allard. 1972. Microgeographical variation in allozyme frequencies in Avena historical events and characteristics which have not been barbata. Proc. Nail. Acad. Sci. 65:2100-2104. considered may partially explain the differences that Hamrick, J.L., Y.B. Linhart, and J.B. Milton. remain. As additional plant species are studied we should 1979. Relationships between life history characteristics and electro- gain a clearer understanding of the role that life history phoretically-detectable genetic variation in plants. Ann. Rev. Ecol. Syst. 10:173-200. characteristics, or combinations of characteristics, play in Hiebert, R.D. shaping the genetic structure of plant populations, 1977. The population biology of bristlecone pine (Pinus longaeva) in including those of forest trees. the eastern Great Basin. Ph.D. dissertation. Univ. of Kansas, Lawrence, 82 p. Jain, S.K., and A.D. Bradshaw. 1966. Evolutionary divergence among adjacent plant populations. I. Acknowledgments: The evidence and its theoretical analysis. Heredity 21:407-441. We wish to acknowledge the technical assistance provided by Diane Küchler, A.W. Bowman, Kareen Sturgeon, and Karen Hamrick throughout the 1964. Potential natural vegetation of the conterminous United States. preparation of this paper. We especially wish to thank those workers who Amer. Geogr. Soc. Spec. Publ. 36. have shared their unpublished data with us: Thomas Adams, M. Lerner, I.M. Thompson Conkle, Robert Eckert, Lauren Fins, Ray Guries, F. Thomas 1954. Genetic homeostasis. 134 p. Oliver and Boyd, London. Ledig, Constance Millar, Richard Morris, Thomas Snyder, Andrew Libby, W.J., R.F. Stettler, and F.W. Sets. Torres, and Francis Yeh. Portions of this research were supported by NSF 1969. Forest genetics and forest tree breeding. Ann. Rev. Genet. 3:469- Grants DEB 76-01295 and BMS 75-14050. 494. Lundkvist, K. 1979. Allozyme frequency distributions in four Swedish populations of Norway spruce (Picea abies K.). 1. Estimates of genetic variation LITERATURE CITED within and among populations, genetic linkage and a mating system parameter. Hereditas 90:127-143. Neale, D.B. Adams, R.P. 1975a. Gene flow versus selection pressure and ancestral differentiation 1978. Allozyme studies in balsam fir. M.S. Thesis, University of New in the composition of species: analysis of populational variation of Hampshire, Durham, N.H. 83 p. Juniperus ashei Buch. using terpenoid data. J. Mol. Evol. 5:177-185. Nevo, E. Adams, R.P. 1978. Genetic variation in natural populations: patterns and theory. 1975b. Numerical-chemosystematic studies of infraspecific variation in Theor. Popul. Biol. 13:121-177. Juniperus pinchotii. Biochem. Syst. Ecol. 3:71-74. Raunkiaer, C. Antonovics, J. 1934. The life forms of plants and statistical plant geography; being the 1971. 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