Botany Publication and Papers Botany

4-1992 Male sterility in soybean and maize: developmental comparisons R. G. Palmer United States Department of Agriculture

M. C. Albertsen Pioneer Hi-bred International

H. T. Horner Iowa State University, [email protected]

H. Skorupska Clemson University

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Recommended Citation Palmer, R. G.; Albertsen, M. C.; Horner, H. T.; and Skorupska, H., "Male sterility in soybean and maize: developmental comparisons" (1992). Botany Publication and Papers. 76. https://lib.dr.iastate.edu/bot_pubs/76

This Article is brought to you for free and open access by the Botany at Iowa State University Digital Repository. It has been accepted for inclusion in Botany Publication and Papers by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. Male sterility in soybean and maize: developmental comparisons

Abstract Sexual reproduction in angiosperms is a complex process that includes a portion of the sporophytic (spore- producing) generation and all of the gametophytic (gamete-producing) generation. The ag metophytic generation includes the developmental stages from the end of meiosis to fertilization. For normal reproduction, coordination of both female and male reproductive ontogenies must occur. An abnormality anywhere in this process may lead to sterility.

Disciplines Agronomy and Crop Sciences | Botany | Plant Breeding and Genetics

Comments This article is published as Palmer, RG, MC Albertsen, HT Horner, and H Skorupska. 1992. Male sterility in soybean and maize: developmental comparisons. The Nucleus 35:1-18.

Rights Works produced by employees of the U.S. Government as part of their official duties are not copyrighted within the U.S. The onc tent of this document is not copyrighted.

This article is available at Iowa State University Digital Repository: https://lib.dr.iastate.edu/bot_pubs/76 the nucleus vol 35(1) 1-18, 1992

MALE STERILITY IN SOYBEAN AND MAIZE DEVELOPMENTAL COMPARISONS '

2 3 4 RG PALMER1, MC ALBERTSEN , HT HORNER , H SKORUPSKA

1Research Geneticist, United States Department ofAgriculture-Agricultural Research Service, Field Crops Research Unit, Professor, Departments of Agronomy and Zoology/Genetics, Iowa State University, Ames, Iowa 50011-1020 USA; 2Geneticist, Biotechnology, Pioneer Hi-Bred International, Johnston, Iowa 50131 USA; 3Professor of Botany and Director of Bessey Microscopy Facility, Iowa State University, Ames, Iowa 50011-1010 USA,· 4Associate Professor, Department of Agronomy and Soils, Clemson University, Clemson, SC 29634-0359 USA

Received on December 26, 1991

Sexual reproduction in angiosperms is a com­ ile, female-sterile mutations (33, 34, 51) or plex process that includes a portion of the male-sterile, female-fertile mutations (18, 32, sporophytic (spore-producing) generation and 44, 45, 48). all of the gametophytic (gamete-producing) generation. The gametophytic generation We have chosen to describe sterility systems includes the developmental stages from the in soybean, Glycine max (L.) Merr., and in end of meiosis to fertilization. For normal maize, Zea mays (L.). These two species arc reproduction, coordination of both female and representative of the Leguminosae and the male reproductive ontogenies must occur. An Gramineae. In addition, these two species arc abnormality anywhere in this process may important crop plants throughout the world. lead to sterility. Our objectives are: 1) to describe the genetics Classification of sterility into various catego­ and development of male-sterile, female-fertile ries has been reported by many investigators. mutations in soybean and in maize and 2) to Gottschalk and Kaul (32) and Johns et al. ( 46) compare and contrast developmental abnor­ divided male sterility into structural (func­ malities among soybean mutants and among tional) and nonstructural. Structural refers to maize mutants. absence of anthers or archesporial tissue, the lack of pollination resulting from indehiscence Soybean : Normal development of anthers of anthers, or spatial separation of male and and pollen female organs. Nonstructural includes all The reproductive biology of the soybean has abnormalities that interrupt microsporogenesis been summarized (12). A brief description is and microgametogenesis. On the basis of necessary to compare development in male­ inheritance patterns, nonstructural sterility may sterile plants. The keel petals of the flowers be classified as cytoplasmic or nuclear. Cyto­ appear first, followed by the wing petals and plasmic male sterility has been reviewed by then the standard petal. While the petal pri­ Duvick (23), Laser and Lcrsten (52), Hanson mordia are small, the stamens are initiated in and Conde (41) and Pring and Lonsdale (66). the upper whorl first, followed by a lower Nuclear male sterility may include male-ster- whorl of five stamens. Each stamen is differ-

Joint contribulion of USDA ARS, Field Crops Research Unit and Journal paper no. J-12983 of The Iowa Agric. and Home Econ. Exp. Stn., Ames, Iowa SOOll - 1010 USA project 2985. 2 RG PALMER ET AL

Table 11 Genetics of male- sterile, female ·fertile mutants in soybean (GT= Genetic type, Collection number)

Mutant GT Source Literature

msl (North Carolina) T260 Farmer's field in 1966 in North Carolina Brim, Young (9)

msl (Tonica) T267 Field of Harosoy by F.M. Bur&ess in 1955 at Palmer et al. (59) Tonica, Illinois

1 msl (Urbana) T266 In an F3 row of L67-533 (Clark x Higan) x Boerma Cooper (8); P.-. SFR 300 in 1971 at Urbana, Illinois et al. (59)

msl (Ames-1) T268 In T258 in 1970 at Ames, Iowa Palmer et al. (59)

msl (Ames-2) T287 In S4,, progeny derived from AP6(Sl) Cl Skorupska, Palmer (75) population in 1984 by Ron Secrist at Ames, Iowa

ms (Sbenong) In L-78-387 in China Yee, Jian (82)

msl (Danbury) T290 In field of Beeson in Danbury, Iowa, outcross, Skorupska, Palmer (75) ~~ male parent unknown .~ ·+ ms2 T259 In F3 pf SLll (Wayne-r Rpm Rpsl x L66L-177 Graybosch et al. (35); ~;(J; [Wayne x (Hawkeye x Lee)] in 1971 at Graybosch, Palmer (36) ;;f~ Eldurado, Illinois ·., :~:',fr msJ (Washington) T273 In F3-plant progeny row of Calland x Cutler in Palmer et al. (60) ·W 1971 at Washington, Iowa

msJ (Flanagan) T284 In field of Wabash in Flanagan, II~ outcross, Chaudhari, Davis (13); male parent unknown Grayboscb, Palmer (38)

msJ (Plainview) T291 In F 2 generation, in cross (Viking x Classic II) Skorupska, Palmer (75) x (Mitchell x Columbus) by William Davis, in Plainview, Tx

ms4 (Ames) T274 Semisterile plant found in Rampage in 1913 at Delannay, Palmer (22) Ames, Iowa

ms4 (Fisher) T292 In field of Corsoy by William Davis, in Fisher, Skorupska, Palmer (75) Arkansas

ms5 T277 Selected at Blacksburg, Virginia in 1976 in M3 Buss (11) generation of neutron-irradiated Esse"

ms6 T295 In line A78-145014 (From A74-204034 X Skorupska, Palmer(74); C1520 in 1978 at Ames, Iowa Palmer, Skorupska (63)

msp T271 In a 40-parent bulk population [AP6(Sl)Cl} in Stelly, Palmer (76,77) 1975 at Ames, Iowa

entiated into an anther and a filament. The gium consists of the following cell types : filaments of nine stamens eventually coalesce, epidermis, endothecium, two or three parietal giving rise to a diadelphous condition. Four layers, a tapetum consisting of uninucleate locules (microsporangia) are differentiated per cells and several rows of sporogenous (male) anther. Preceding meiosis, each microsporan- cells. Connective parenchymatous cells, many MALE STERILITY 3

Table 2: Sourct!S of cytoplasms for cytoplasmic male - tapetal cells remain uninucleate throughout sterility in soybean • anther development Upon completion of meiosis, enlargement of the endothecium Adelphia Elf Magna commences. Chippewa Ford Prize Microgametogenesis is characterized by at Chippewa 64 Grant Provar least five stages. The four microspores in a quartet are arranged tetrahedrally within a Clark Haro50y Rampage callose sheath and are somewhat angular in Oassic I Harosoy 63 Shelby appearance. Pollen-wall formation is initiated at this time and upon callose dissolution, the Classic II Hobbit Traverse microspores become more spherical. The Columbus Kent Wayne microspores enlarge and contain many small vacuoles. At this early vacuolate microspore Cutler Lindarin Wirth stage, microspore wall formation continues Disoy Lindarin 63 Williams with the development of the individual layers : tectum, columellae and endexine. Three colpi • W.H. Davis (1985) Route to hybrid soybean are evident. The microspore cytoplasm is rich production, United States Patent 4,545,146. in starchless plastids and small vacuoles. At the late vacuolate microspore stage, vacuoles expand, the tapetum begins to degenerate and containing calcium oxalate crystals, hold the the endothecial layer expands. Endothecial Iocules together. A single vascular strand expansion results from the deposition of U­ traverses this latter tissue (10). shaped thickenings on the radial and inner tan­ gential walls. Microsporogenesis is characterized by the sporogenous and meiotic stages (fig 1). The primary sporogenous cells mature into two or Table 3: Sourct!S of rt!Storer genes for cytoplasmic male three rows of microspore mother cells sterility in soybean • (MMC). Numerous plasmodesmata connect the sporogenous cells to each other and to the surrounding tapetal cells. rl rl source Bedford Dyer Kirby The first meiotic division is highly synchro­ nous within a given microsporangium, where­ Bethel Forrest Tracy as less synchrony exists among MMCs during Centelinial Hill Wabash the second meiotic division. As the MMCs enter meiosis, callose secreted interior to the Dare York primary cell wall severs all connections be­ r2 r2 source tween adjacent cells. Cytokinesis does not occur until the completion of the second Bragg Hardee Major meiotic division. A synchronous furrowing Braxton Hutton Ogden takes place, dividing the four-nucleate cell into four haploid microspores. During meiosis, Cobb Jackson Volstate the tapetal cells become cytoplasmically dense Govan Kirby Wright and rich in organelles. The radial and inner •w. H. Davis (1985) Route to hybrid soybean production. tangential tapetal walls partially dissolve as United States Patent 5,455,146. meiosis proceeds to the quartet stage. The 4 RG

Microspore mitosis gives rise to a vegetative grains. A matrix of unknown origin preventa r-?­ cell and a smaller generative cell. Pollen wall the release of CMs from anthers. Coenocytic ;~ formation continues in the vacuolate pollen microspores differentiate colpi, but the number j · with the production of the intine. At the en­ and distribution of these structures are vari- ::i, gorged pollen stage, intine formation is com­ able. Pollen tubes are generated rarely by pleted and starch and lipid bodies are evident. CMs both in vitro and in vivo. It remains Tapetal degeneration is completed, parietal unclear if these pollen tubes release sperm and ,~ layers are crushed and the endothecium is ex­ effect fertilization (3, 15, 72). Probably some ,; panded fully. gametes from coenocytic microspores are able , to participate in fertilization at low frequency, Male-sterile female-fertile mutations in or apomixis occurs in msl msl plants (15). soybean The Soybean Genetic Type Collection is part In ms] msl male-sterile plants, the absence of of the USDA Soybean Germ plasm Collection postmeiotic cytokinesis is the only observable and is composed of strains used in qualitative phenotypic abnormality. The ms4 male-sterile genetic studies. Table 1 lists the male-sterile, mutation influences the process of postrneiotie female-fertile mutants by gene symbol, Type cytokinesis as well. Cell plates frequently are number, origin and relevant literature on the incomplete or irregular in orientation. The genetics. Recent information on the genetics nature of postmeiotic cytokinesis in reproduc­ and cytology of sterility mutations in soybean tive cells is variable. Some anthers (3.3% of are available (39, 61, 62). the anther loci examined) undergo complete and properly oriented cytokinesis (22, 37). Cytoplasmic male sterility and restorer genes The process is influenced by temperature. The have been reported only once in soybean (21). occurrence of several independent mutations at Sources of cytoplasm and restorer genes are the msl and ms4 loci raises questions about given in tables 2 and 3, respectively. These the sensitivity responsible for post-meiotic gene symbols have not been submitted to the events and microspore division (75). Soybean Genetics Committee. Developmental reproductive patterns in male-sterile, female­ The lack of normal cytokinesis associated with fertile soybean mutations have been summa­ ms] and ms4 genes might be correlated with rized (39). Reported male-sterile, female­ aberrant cytoskeletal differentiation. Jn addi­ fertile mutations do not suppress the gross tion, the ms4 mutation influences abnormal differentiation of anthers. However, such pollen wall differentiation of CMs, which mutations, in the homozygous recessive condi­ results in the formation of 1 to 4-celled aggre­ tion, produce mostly nonviable pollen grains. gates that eventually degenerate (22), The There are significant differences between the abnormalities of pollen-wall formation may be action of male-sterile genes at msl, ms2, ms3, indicative of malfunction of the plasmalemma. ms4 and ms6 loci and, to some extent, some similarities (table 4). Mutations at the msl Tapetum malfunctions in anthers are due to locus induce male sterility by acting on the the action of ms2, ms3, ms6 and msp loci function of the reproductive cells alone. Nu­ (table 4). In male-sterile plants, the quartet clear division within the meiocytes proceeds stage initially seems similar to that of male­ in a typical fashion in msl msl plants. How­ fertile plants. Cytokinesis occurs, but micros­ ever, the absence of cytokinesis results in the pores degenerate after organizing probaculae, production of quadrinucleate (coenocytic) the primordial sporopollenin deposits charac­ microspores (CM). These cells produce walls teristic of pollen walls. Callose dissolution and of sporopollenin and engorge with reserve microspore-wall formation are not observed. materials in the same fashion as viable pollen Ultrastructural observations indicate that MALE STERILITY 5

T•ble 4: Phenotypic expression of male- sterile, female-fertile mutants in soybean

Mutant Developmental abnormalities Relevant literature

ms] All msl mutants have normal meiosis but male reproductive cells have Albertsen, Palmer failure of cytolc.inesis following telophase II, resulting in coenocytic (3); Kennell, Horner microspores. The T266 mutant has greater seed set than the other mutants (49); Kenworthy et al tested. Female abnormalities are less in T266 than in the other mutants. (50); Beversdorf, Abnormalities include polyembryony and polyploidy among both mooo­ Bingham (7); Chen et embryonic and polyembryonic seeds. al (16); Cutter, Bingham (20)

ms2 Tapetal cells form abnormally large vacuoles during prophase I. Male Graybosch et al (35); sterility is due to an abortion of quartets. Callose dissolution and initiation of Graybosch, Palmer microspore walls do not occur. (36)

ms3 Abnormalities of tapetal cells sometimes occur before and sometimes after Palmer et al.(59); microspore degeneration. Male sterility is due to abortion of microspores. Nalc.ashima etal.(55) ; Callose dissolution does not occur. Microspore wall formation initiated but Buntman, Horner not completed. (10); Graybosch, Palmer (38)

ms4 Cytolc.inesis following telophase II may be absent, incomplete or disoriented, Delannay, Palmer resulting in cells with different numbers of nuclei. Coenocytic microspores (22); Graybosch, are produced. No abnormalities in female reproduction have been reported. Palmer (37)

ms6 Proper meiosis leads to normal telophase II. Tapetal behavior is affected Slc.orupslc.a, Palmer developmentally and all layers of the anther wall were intact at anthesis. (74) Microspores degenerated at quartet stage. Sterile plants have smaller flowers and shrunlc.en anthers.

msp Effect on microsporogenesis and microgametogenesis is inconsistent. Stelly, Palmer (78) Abortion of cells may occur at any time between late sporogenous and pollen stages. In tapetal cells, unusually large vacuoles generally precede abnormalities in tapetal cells.

tapetal cells fonn abnonnally large vacuoles and proteins and a single-staining procedure during prophase 1 of meiocyte meiosis and for sporopollenin to find developmental de­ that failure of the tapetal layer to differentiate tectable features of the ms3 mutation. They properly is the cause of male sterility. The concluded that, in ms3 ms3 plants, abnonnali­ mature tapetum is represented by cells each ties are expressed during early anther develop­ consisting of a large central vacuole, little ment, probably beginning at meiosis, that di­ cytoplasm, and a few organelles. The inner rectly influence the malfunctioning of the tangential cell wall fails to dissolve and the tapetum. At the quartet stage, tapetal cells tapetum collapses along with microspores seem normal. As microspore degeneration (35). In ms3 ms3 plants, tapetal degeneration occurs, however, tapetal cells either collapse may precede or follow microspore degenera­ or accumulate the material that is a precursor tion (38). In the msp msp genotype, tapetal or modified fonn of sporopollenin. Because of abnonnalities (premature vacuolation, cellular the premature degeneration of the ms3 ms3 collapse) precede the abortion of reproductive tapetum this material, which fluoresces in UV cells (78). Nakashima et al. (55) used a triple­ light, is not transferred to the locules and staining procedure for DNA, polysaccharides, microspores and as a result, accumulates in 6

Table 5: Gendic test for origin of embryo and endosperm in soybean (Chr. No. = Chromosome number)

Generation Genotype Phenotype

Chr. No. Flower Plant color Fertility color

Female wl wl Yll yll msl 2n=2x=40 white yellow- green sterile msl

Male WI WJ WI WI Yll 2n=4x=80 purple green fertile YJJ Yll Yll Msl Msl Msl Msl

F1 WJ WI wl wl Yll Yll 80 purple green fertile YJJ Yll Msl Msl msl msl

F1 WI WI wl wl Yll Yll 80 purple yellow- green fertile yll yll Msl Msl msl ms]

F1 WI WI wl wl Yll Yll 80 purple green- yellow fertile Yl J yll Msl Msl ms] msl

the tapetum. The thinness of the ms3 ms3 stage. Therefore, the association between microspore walls supports this observation. callose dissolution and tapetum might be an Pollen-wall structure differs from that of additional factor causing male sterility. male-fertile plants in the omission of the columellar layer. The intracellular transport Mutation at the ms6 locus affected the proper from the tapetum to the microspore and/or differentiation and function of the primary conversely, the inability of the microspore to tapetal layer. In male-sterile ms6 ms6 plants, assimilate and metabolize sporopollenin pre­ all layers of the anther wall including the cursors may be blocked by the ms3 ms3 epidermis, endothecium, two parietal layers, system. Less extensive deposits of sporopolle­ expressed cell hypertrophy and degenerated nin were noted within the tapetum and anther tapetal cells were present in mature anthers. locules of ms4 ms4 sterile plants (37). Subsequently, tapetum malfunction leads to microspore degeneration at the quartet stage In both mutations, ms2 and ms3, failure of (74). callose to break down at the proper time was noticed. In ms2 ms2 sterile plants, mature Our studies on male-sterile,' female-fertile anthers contain degenerate microspores and mutations in soybean support numerous other callose fails to dissolve (36). In ms3 ms3 reports on both genetic and cytoplasmic male sterile plants, callose breakdown occurs preco­ sterility implicating the tapetum as the primary ciously and the microspores are released too tissue responsible for male sterility with the early. Warmke and Overman (79) reported exception of the msl and ms4 mutations. differences in callose formation hctween male­ fertile and cytoplasmic male-sterile sorghum Megagametogenesis in msl sterility mu­ anthers. Frankel et al. (28) reported that cyto­ tations in soybean plasmic male sterility in Petunia was due to The msl system displays pleiotropic effects on failure of callose dissolution at the proper the female reproductive system. The msl MALE STERILI1Y 7

Table 6: Genetic test for apomixis on msl msl soybean plants in F2 generation.

Genotype Phenotype

Flower color wl wl Yll yll Msl ms] Plant color Fertility self-pollination l white yellow- green fertile

1•• wl wl Yll Yll Msl ms] white green fertile

2 wl wl Yll Yll Msl ms] white green fertile

wl wl Yll Yll ms] ms] white green sterile

2 wl wl Yll yll Msl Msl white yellow- green fertile

4 wl wl YJJ yll Msl ms] white yellow- green fertile

2 wl wl Yll yll ms] msl' white yellow- green sterile

wl wl yll yll Msl Msl white yellow fertile

2 wl wl yll yll Msl ms] white yellow fertile

wl wl yl I yll ms] msl white yellow sterile

• Desired genotype; •• Genotypic frequency

gene is associated with reduced female fertili­ extent of postmeiotic callose wall formation; ty and with increased frequencies of polyem­ the degree of nuclear fusion occurring in bryony, polyploidy and haploidy in the proge­ megasporogenesis and early megagameto­ ny of sterile plants (9, 16, 50). genesis where post meiotic callose walls do not form, and the survival of each nucleus Cytological investigations of ovules from msl during megagametogenesis. (iv) The numbers msl plants have uncovered abnormal megaga­ of the gametophytic cells were inconsistent, metophytes that contain supernumerary nuclei yet the positioning of the antipodals, central (19, 20). Kennell and Homer (49) studied the cell, polar nuclei, and egg apparatuses usually development of the female gametophytes of was unaffected. (v) Multiple cells, up to four msl msl plants. They reported that : (i) Com­ egg apparatuses, were present at the micropy­ plete or partial absence of postmeiotic cytoki­ lar region of the mature megagametophyte. nesis callose walls could lead to a coeno­ megaspore of four haploid nuclei. Ultrastruc­ The msl mutation does not have complete tural examination of re-embedded 1 µm sec­ female fertility. Some female sterility occurs, tions showed a lack of dictyosomes and their and msl msl plants may have megaspore wall associated vesicles that were present in a formation following meiosis or incomplete or fertile megaspore after chromosome segrega­ complete failure of cytokinesis, depending tion at meiosis 1. (ii) The number of mitotic upon the source of msl and the environment. divisions of megaspores did not seem to be Double fertilization with pollen from a male­ affected by the msl gene. (iii) The number of fertile diploid can result in the production of nuclei in megagametophytes seemed to be a normal primary endosperm in the presence determined by three factors, namely, the of a zygote of independent ploidy level. This 8 RG PALMER ET AL

Table 7: Possible use of tetraploid soybean plants with Yll yll, is inherited as 1 green (Yll Yll) : 2 the msl allele in crosses with perennial Glycine species yellow-green (Yll yll) : 1 yellow (yll yll) to scale down the ploidy level. plants. Crosses were conducted between yel­ low-green male-sterile plants (msl msl Yl 1 Female parent Male parent" (2n=4x=80 yll) and green fertile tetraploid cultivars (Msl (2n=4x=80 chromosomes) Msl Msl Msl Yll Yll Yll Yll). We would chromosomes) expect 80-cbromosome F1 plants because the selection process would be for 40-chromo­ Msl Msl Msl Msl Msl Msl Msl some eggs, or cells that can function as eggs, msl to be fertilized by 40-chromosome pollen

or Msl Msl msl (table 5). These 80-chromosome F1 plants msl should have the correct endosperm-embryo relationship. Other possibilities include 20- or Msl msl msl chromosome plants (maternal haploid), 40- msl chromosome plants(patemal haploid) or 60- or msl msl ms] msl chromosome plants. Sixty-chromosome plants have been obtained from msl msl plants F1 Msl Msl grown in the field in the absence of 80-chro­ mosome plants. The 60-chromosome plants F2 ms] msl msl msl" were either msl msl msl or Msl msl msl but • Perennial Glycine species; never Msl Msl msl (14). Among the 80- •• These plants might be capable of producing 40- chromosome plants, the expected results are chromosome embryos via the mechanism(s) that give (table 5) : (i) If the seed arises from duplica­ 20-chromosome embryos from 40-chromosome msl tion of egg cells or some other gametophyte msl plants. cells, the F 1 seedling color would be either green (Yll Yll Yll Yll) or yellow-green (Yll Yll yll yll); (ii) If the seed arises from somatic cells, the F1 seedling color would be mechanism permits haploidy and polyploidy green-yellow (Yl 1 YI I Yll yl l). These geno- from aberrant megagametophytes. Table 8: Possible use of tetraploids with the msl in Application of msl sterility mutations in crosses with deficiency aneuploids at the tetraploid level soybean to produce deficiency aneuploids at the diploid level !')Embryo-endosperm relationships : The success of the embryo depends upon the Female parent (78 Male parent (2n=4x=80 normal development of the endosperm in chromosomes) chromosomes) almost all species. In previous studies, attemp­ ts to obtain triploid soy~ean plants (2n =3x = Msl Msl Msl Msl Msl Msl Msl Msl or 60) through natural cross-pollination and Msl Msl msl msl or artificial cross-pollination between autotetra­ Msl msl ms] msl ploids and diploids or reciprocal crosses, were F Msl Msl __ 79 chromosomes unsuccessful (69). These results might be 1 explained by endosperm-embryo ploidy imbal­ F2 ms] ms] msl msl" 78, 79, 80 chromosomes ance. The male-sterile mutant msl msl was used in studies to determine the embryo-endo­ • These plants might be capable of producing 38, 39, or sperm relationships in soybean. 40 - chromosome embryos via the mechanism(s) that give 20 - chromosome embryos from 40-chromosome msl msl plants. The codominant chlorophyll-deficient mutant, MALE SIBRILITY 9 types can be distinguished from each other by ing. The pollen is stained with 12 Kl, and the plant color, pigment determination measured fertile plants are removed. The desired geno­ spectrophotometrically or by genetic segre­ type is wl wl Yll yll msl msl, and these gation of the yll allele(s) in the F2 generation plants are grown in isolation from other soy­ (43). bean plants in the greenhouse (table 6). Seeds are harvested from these plants. Zhang and Palmer (83) reported that the segregation ratio among the F1 plants was Expected results are that all plants will be close to the expected 1 green : 2 green-yellow white flowered and male sterile. Chromosome : 1 yellow-green. They concluded that : (i) number of all plants will be determined. Plant triploids were not produced; (ii) tetraploid color is the phenotype that will help us deter­ progeny can be produced by the fusion of 2n mine the origin of the seed. Expected results msl eggs or fusion of other Zn gametophyte are (table 6) : (i) If the seeds arise through cells in the embryo sac with a 2x sperm from functional pollen (should be infrequent), the tetraploid plants; (iii) the megaspore mother seedling color will be in a ratio of 1 green : 2 cell of msl msl plants undergoes meiotic yellow-green : 1 yellow; (ii) If the seeds arise division without cytokinesis after telophase II from somatic cells, seedling color should be and forms more than the normal number of yellow-green only; (iii) If the seeds arise from gametes, which can fuse with each other to egg cells or some other gametophyte cells, generate tetraploid gametophyte cells. seedling color will be either green or yellow and seedlings will be haploid chromosome 2) Test for apomixis : An experiment has been number; (iv) If the seeds arise from egg cells designed to test for apomixis by using msl or some other gametophyte cells that are msl soybean plants and wl and yll marker duplicated or fused, seedling color will be alleles (table 6). The chlorophyll-deficient either green or yellow and seedling will have mutant Yl 1yl1 has a yellow-green phenotype, diploid chromosome number. whereas Yll Yll plants are green and yll yll plants are yellow lethal. Purple-flower plants 3) Scaling down the ploidy level : Eighty­ can be WJ WJ or Wl wl and white-flower chromosome plants have been identified plants are wl wl. A few seeds are found on among progeny of ms] msl plants. The geno­ ms] ms] plants grown in the field or in the type of these plants was either msl msl msl greenhouse. One seed has been observed from msl or Msl msl msl msl (16). We have 430 emasculated flowers from msl msl plants found 40-chromosome plants (msl msl) grown in the greenhouse in isolation from among progeny of 80-chromosome ms] msl fertile plants. This seed gave rise to a male­ msl msl plants. In table 7, we suggest using sterile, 40-chromosome plant (15). We believe 80-chromosome plants with the msl allele in that this seed was not the result of self-polli­ crosses with perennial Glycine species to scale nation or natural cross-pollination. We hy­ down the ploidy level to the 40-chromosome pothesize that this seed was the result of number. We have observed 78 and 79-chro­ apomixis. We are uncertain whether the seed mosome plants among progeny of 80-chromo­ was the result of a fusion event or the result some Msl Msl Msl Msl soybean plants. The of a diploid cell functioning as an embryo. 78-chromosome plants have been true breed­ ing for 78-chromosome plants for several All the plants should be white flowered (table generations. Deficiency aneuploids at the 6). The green plants are identified as seedlings diploid chromosome level have been reported and removed and the yellow plants are lethal. (73). In table 8, we suggest using plants with The fertile plants and sterile plants cannot be the msl allele to scale down the ploidy level distinguished from one another until flower- of 78 and 79-chromosome plants to give 38 10 RG PALMER ET AL

Table 9: Phenotypic expression of male-sterile, female-fertile mutants in maize (CN = Chromosome number) conli

Mutant CN Developmental abnormalities Relevant literature

msl 6 Microspores seem to develop dark thickened walls; some Singleton, Jones (70); precociously condensed chromosomes. Breakdown at Beadle ( 6); Albertsen, midvacuolate microspore stage. Phillips (4)

ms2 9 Lack of significant microspore wall development; some Eyster (25); Albertsen, precociously ci>ndensed chromosomes. Breakdown at Phillips (4) midvacuolate microspore stage.

ms3 3 Breakdown by midvacuolate microspore stage. Eyster (26)

ms5 5 Normal microspore development until microspore mitosis Beadle (6); Albertsen, stage when further development is arrested. Phillips (4)

ms7 7 Similar to ms2. Beadle (6); Albertsen, Phillips (4); Morton et 11L (56)

ms8 8 Meiosis may or may not be completed. Any microspores Beadle (6); Albertsen, that form degenerate. Phillips (4)

ms9 Similar to ms8. Beadle (6); Albertsen, Phillips (4)

msJO 10 Microspore wall development lags behind microspore Beadle (6); Albertsen, vacuolation. Breakdown at midvacuolate microspore stage. Phillips (4)

msll Normal microspore development until microspore mitosis Beadle (6); Albertsen, stage when further development is arrested. Phillips (4)

msl2 Normal microspore wall development but arrested nuclear Beadle (6); Albertsen, development. Breakdown at midvacuolate microspore stage. Phillips (4)

and 39-chromosome plants. develop into three anthers, each with a fila­ ment. One anther becomes larger than the Maize : Normal development of anthers other two anthers. Four locules (microsporan­ and pollen gia) are formed per anther. Before meiosis, The reproductive biology of maize bas been each microsporangium .consists of the fol­ summarized (4, 17, 68). The term microsporo­ lowing tissues : epidermis, endothecium, genesis often is used to describe the entire usually one parietal layer, a tapetum com­ process of sporogenesis and gametogenesis in posed of uninucleate cells and the sporoge­ the development of mature pollen grains (fig nous (male) cells. Connective tissue holds the 2). A brief description is necessary to compare four locules together. The single vascular development between male-fertile and male­ strand runs through this last tissue. Micro­ sterile plants. Male flowers (florets) develop sporogenesis is characterized by the sporo­ in a terminal inflorescence called a tassel. genesis and meiotic stages (fig 2). The prima­ Each floret is enclosed by two bracts, the ry sporogenous cells are pie-shaped and are lemma and palea. Inside, stamen primordia separated by primary walls with interconnect- MALE SlERILITY 11

Table 9 (contd.):Phenotypic expression of male-sterile, female-fertile mutants in maize (chromosome number= CN)

Mutant Chromo­ Developmental abnormalities Relevant literature some

ms13 5 Microspore wall development lags behind microspore Beadle (6); Albertsen, vacuolation; some precociously condensed Phillips (4) chromosomes. Breakdown at midvacuolate microspore stage.

msl4 Normal microspore development until late vacuolate Beadle (6); Albertsen, microspore stage. Phillips (4)

msl7 Abnormalities observed at any stage after meiotic Emerson (24); Albertsen, prophase. Spindle development and orientation Phillips (4) commonly affected.

ms20 (9)? Breakdown by midvacuolate microspore stage. Eyster (27); Albertsen (unp)

ms22 MMCs degenerate before pachynema. West, Albertsen (81)

ms23 3 Degeneration begins during meiosis I but nuclear West, Albertsen (81); events continue until meiosis II . Degeneration by Albertsen (1) midvacuolate microspore stage

ms24 Very late pollen breakdown; irregular starch West,Albertsen (81) accumulation from barely detachable to fully engorged

ms28 Anaphase I disturbed Golubovskaya (30)

Ms41 4 Dominant male sterile. Neuffer et al. (57)

ms43 8 Anaphase I irregular; functional disturbance of spindle Golubovskaya, Distanova apparatus. (31)

Ms44 4 Dominant male sterile; breakdown occurs by late Golubovskaya (30); vacuolate stage. Albertsen, Sellner (2); Albertsen, Neuffer (5)

ing plasmodesmata to each other and to the Callose wall fonnation following meiosis tapetum. Preceding meiosis, callose walls are and mehsis II always occurs so that all the formed interior to the primary walls surround­ microspores formed are adjacent to the tape­ ing the microspore mother cells (MMCs). tum. The area of each microspore in a quartet MMCs subsequently round up and sever their that is adjacent to the tapetum is the position plasmodesmata with all adjacent cells. Each where a single pore will fonn. Before the cal­ MMC remains adjacent to the tapetum. The lose is digested away to allow separation of MMCs enlarge and pass through meiosis I. A the microspores, a young pollen wall (com­ callose wall is formed between the two result­ posed of sporopollenin) is initiated around ing nuclei of a meiocyte, creating dyads. After each microspore, along with a single pore. meiosis II, two more callose walls fonn, which delineate a quarter of microspores. The majority of tapetal cells undergo nuclear 12 RG PALMER ET AL m•2 m•s degeneraUon bl tetrads

t ms3 degeneraUon of mk:rotpen1 / \

sporogenous cell

(2N) vacuole pol~n WIU forms thickens msp colpus...... , degeneration coenocytlc of cells at micro spore any stage

generative engorged cell vacuolate pollen pollen

{@/ '~~~ /'-""'\nofmicrospores :< (:j}. 1 meiosis! ? ~ meiosls ll ~==!~===! ms, \ . \;W1 .· / cell wall \ \.. {;)Q) J cell wall r, \ / forms \~/ formation ,o"~ , ..., m•22 ~o"" microspore dyad Cf mother cell microspores @ non-vacuolate sporogenous micros pore cell (N) m•12 (2N) m•10, 13 vacuole pollen wall forms lthickens degeneration of cells at any stage ~e-·

vacuolate microspore

sperm cells

pore engorged vacuolate pollen pollen MALE STERILITY 13 division without cytokinesis to produce binu­ Male-sterile, female-fertile mutations in cleate cells. Maize tapetum, like several other maize species studied, does not become wholly binu­ Table 9 gives a summary of 21 nuclear male­ cleate during any stage of its development. sterile mutants in maize. Other nuclear male However, the greatest increase in binucleate steriles have been described in the literature, cells takes place during the meiotic period. An but either stocks are no longer available or ge­ orbicular wall (composed of sporopollenin) netic tests showed allelism. We have not develops on the inner tangential surface of the attempted to determine the number of instanc­ tapetum next to the microspores. The outer es in which independent mutations of a given tangential and radial primary walls around male sterile have occurred. For some loci, each tapetal cell partially dissolve. In many there have been many. At least 20 additional places, they seem to be absent. Each tapetal nuclear male steriles currently are un­ cell cytoplasm is dense and rich with organ­ described. They are being test crossed and elles during the later meiotic stages. described (EB Patterson, Univ. of Illinois, pers. commun.; MC AJbertsen, pers. com­ After callose digestion, each microspore and mun.). We have included only mutants that fit its single pore remain adjacent to the tapetum the strict definition of a male-sterile mutant, as the microspore enlarges. At this time, the that is, the mutation affects male fertility and microspores have dense cytoplasms with many bas little or no effect on female fertility. small vacuoles. The microspore walls and Male-sterile mutations in maize are expressed pores continue to become more prominent throughout microsporogenesis. AJl but two are because of sporopollenin deposition from the recessive. The mutant with the earliest effect tapetum. The microspores increase in size and is ms22; the latest is ms24. Expression of become more vacuolate. ms22 occurs before meiosis. Microspore mother cells (MMCs) are fonned, but they As each microspore becomes highly vacuolate degenerate before pacbynema. Expression of and the microspore wall reaches a maximum ms24 occurs after generative cell formation. thickness, the single nucleus is displaced from Starch engorgement may or may not occur. its central position to the periphery. Micro­ Homozygous recessive ms24 ms24 plants can spore mitosis occurs, producing a pollen grain produce limited viable pollen in some inbred with a vegetative cell and a smaller generative backgrounds.The other male-sterile mutants cell. Intine formation begins. The tapetum affect microsporogenesis at characteristic completely breaks down followed by the phases of development. Mutant ms] develops accumulation of starch and other reserves. The abnormally thickened microspore walls. Wall generative cell in each pollen grain divides to thicknesses vary, but in most instances, walls form two sperm cells, followed by complete develop to about twice the thickness of a engorgement and disappearance of the pollen normal microspore wall. This abnormal devel­ vacuole. opment occurs soon after microspores release from quartets. AJthough chromosomes may The mature three-celled pollen grains remain become condensed, no further mitotic stages adjacent to the tapetal orbicular wall. The are observed. Cytoplasmic and nuclear degen­ anther endothecium becomes fully expanded eration were evident after chromosome con­ and the parietal layer collapses, leading to densation and abnormal wall development. pollen dehiscence. Male steriles ms5 ms5, msl 1 msl 1 and ms14

Fig 1: Microsporogenesis and microgametogenesis in normal soybean : timing of abnormalities associated with each male-sterile mutant is depicted by gene symbol. Fig 2: Microsporogenesis and microgametogenesis in normal maize : timing of abnormalities associated with each male-sterile mutant is depicted by gene symbol. 14 RG PALMER ET AL

ms14 have a late effect on microgametogene­ cytologically, but not genetically, similar to sis, although not as late as ms24 ms24. These ms8. Mutant ms28 ms28 is described as exhib­ male steriles degenerate during the microspore iting disruption of anaphase I. mitosis stage. Wall development and nuclear development proceed normally, with micro­ Two dominant male steriles have been de· spore vacuolation occurring as expected. scribed. One has been given the designation Microspore nuclear chromatin condenses Ms41; the other has been designated Ms44. during early phases of the microspore mitosis Both these mutants have been located on the stage, but no later stages of microspore mito­ Jong arm of chromosome 4 and have been sis were evident. shown not to be allelic. Mutant Ms44 shows Male-sterile mutants ms8 ms8 and ms9 ms9 the breakdown of microspores at the late result in abnormal MMCs that exhibit nearly vacuolate stage. The common feature of all normal nuclear development but abnormal these mutations is that they cause male sterili­ cellular development. MMCs appear smaller ty. Although some mutants act similarly, moot that normal and have poorly defined cell of them display a characteristic timing of male boundaries. Although there was variability as cell degeneration. These mutants represent to bow long these MMCs would exist, degen­ some of the critical genes in the pathway eration occurred fairly quickly. affecting some aspect of microsporogenesis. Mutants mslO mslO and ms13 ms13 are simi­ This could include differentiation of other lar in that the microspore wall development cells in the anther, in addition to the devel­ lags behind comparable stages in microspore oping pollen grains, most notably tapetal cells. vacuolation. Microspore pore development is Albertsen and Phillips (4) refer to the tapetal not complete. Degeneration occurs by the layer in their summary of the effects of cer­ midvacuolate microspore stage. Almost com­ tain male-sterile mutants, but critical evalua­ plete wall development occurs in the ms12 tion of the tapetum was not done in their ms12 mutant, despite nuclear degradation study because it was based on anther squash within the microspores. Vacuolation proceeds preparations. Cheng et al. (17) conducted a normally up until the midvacuolate stage, comparison of anther development in mslO when the microspores degenerate. mslO mutant and fertile plants by using light Mutant ms17 ms17 exhibits variable expres­ and scanning electron microscopy. They found sion during microsporogenesis. Spindle forma­ that there were no developmental differences tion often is affected, resulting in chromo­ between the two lines until after the quartet somes scattered throughout the MMC. Abnor­ stage, when the microspores were separate malities can occur at any stage after meiotic from each other. At this time, the mslO mslO prophase. Abnormal cytokinesis results in a homozygote tapetal cells became vacuolate variety of postmeiotic structures, including and the cytoplasm disintegrated. The micro­ coenocytic quartets and structures with greater spores, however, continued to develop to the or fewer than four microspores. The other midvacuolate stage. By the young pollen male-sterile mutants listed in table 9 also can stage, the tapetum was highly disorganized, as be compared with the preceding mutants. well as the parietal layer and the endothecium. Mutant ms3 ms3 bas not been cytologically The endothecium did not produce any wall described, but we estimate its breakdown to thickenings. occur by the midvacuolate microspore stage. Mutant ms20 ms20 is being studied (MC Cytoplasmic male-sterility in maize Albertsen, pers. commun.). Degeneration is Maize has another nuclear system that can estimated to occur at about the midvacuolate undergo mutation to generate male sterility in microspore stage. Mutants ms23 ms23 and addition to that resulting from mutation in the ms43 ms43 have been described as being nuclear genome. Specific rearrangements in MALE SIBRIUTY 15 the mitochondrial DNA have led to three five subgroups consisting of BID, CA, LBN, types of cytoplasmic male steriles (CMS) in ME and S (71 ). These subgroups have been maize ( 42). These have been identified as T identified on the basis of a variable extent of (Texas), C (Charrua) and S (USDA) types of fertility restoration in a series of nuclear back­ cytoplasmic male sterility. Two of these, T­ grounds and by restriction endonuclease CMS and C-CMS, are sporophytic-type patterns. All the subgroups of CMS-S have steriles. The other, S-CMS, is a gametophytic gametophytic restoration of fertility. That is, sterile. In S-CMS, microspores usually devel­ restoration is determined by the nuclear geno­ op to the trinucleate stage before they degen­ type of the pollen grain. In tassels of CMS-S erate. In T and C-cytoplasm plants, micro­ plants heterozygous for restorer genes, pollen spores degenerate by the midvacuolate stage. grains carrying Rf alleles develop normally, Ohmasa et al. (58) examined the tapetal layers whereas pollen grains carrying rf alleles do of these male steriles and determined that the not. In addition to gametophytic restoration, tapetum of N (normal) and S-cytoplasm plants the CMS-S group is characterized by having degenerated when microspores were at the considerable quantities of two plasmid-like uninucleate through binucleate stages. Lee et DNA's in their mitochondria. This group is al. (54) also investigated S-cytoplasm plants. the only cytoplasmic male sterile in which the In T and C-cytoplasm plants, the tapetum chloroplast DNA can be distinguished from remained intact until the time when N-cyto­ that of normal maize by restriction endonucle­ plasm plants flowered. This does not agree ase analysis (64). with results of Lee et al. (53) and Warmke and Lee (80), who concluded that the tapetum Considerable molecular information is avail­ degenerated much earlier. able for the cytoplasmic male steriles. Mito­ chondria from T and C-cytoplasms are charac­ Each of the CMS lines can be distinguished terized by the synthesis of a single additional by response to restorer genes and by restric­ or variant, polypeptide not found in N or S tion patterns from mitochondrial (mt) DNA cytoplasms. In T, this is a 13,000-MW protein (65). The C-cytoplasm sterility, for example, (40); in C, this is a 16,000-MW protein (29). can be subdivided into five subtypes, identi­ In C-cytoplasm, a 15,000-MW protein is not fied as BB, ES, PR, RB and C (67). Although produced, but a 16,000-MW protein is pro­ restriction patterns of mtDNA are quite simi­ duced in its place. The finding that the trans­ lar, each of the five can be distinguished by lation products of S-mitochondria do not molecular evidence of mitochondrial DNA se­ differ significantly from those of N-mitochon­ quence duplications, recombinations and dele­ dria has been given as an indication that there tions; by the population of mitochondrial is a clear distinction between this CMS and minicircle DNA'S; and by mitochondrial DNA the other CMS types. restriction patterns. Each member of the C­ group carries a 1,913-base-pair (bp) mtDNA Summary minicircle, whereas all entries but RB carry a Descriptions of the developmental abnonnalities in male­ 1,445-bp mtDNA minicircle. The other sub­ sterile, female-fertile mutations in soybean and maize have been presented. Specific timing of mutations can groups are distinguished on the basis of re­ have similar or different influence on the sterility in striction endonuclease patterns and hybridiza­ Glycine and Zea. In maize, a mutation at the ms22 locus tion with specific sequence probes. In addi­ bas the earliest effect; MMCs degenerate before pachy­ tion, Kalman et al. (47) concluded that RB nema. Maize mutations ms8, ms9, ms23, ms28 and ms43 exhibits a later degeneration than the usual C­ cause abnormalities, including MMC degeneration, as early as meiosis I. Female fertility is unaffected or only type. slightly reduced by these mutations.

The S-cytoplasm also can be subdivided into In soybean, the male-sterile, female-fertile mutation at the 16 msl locus affects cylokinesis durin& sporogenesis of both female development. Some nuclear male-sterile mutufl the male and female development. In different eenetic operate like many cyloplasmic male-sterile mutants in thll backgrounds, this mutation can reduce female fertility. the tapetum is adversely affected. The tapetum may The mutation at the ms4 locus, which is similar to that of degenerate prematurely or persist until pollen dehiscence. the msl locus, affects cytokinesis following telophase II; In either instance, incorrect timing of tapetal degeneratio1 cytokinesis may be absent, incomplete or disoriented. No may lead lo male sterility. abnormalities in female fertility have been reported for the msilocus. All these studies suggest that a variety of mechanisms associated with male sterility are present in plants. Eveii No male-sterile, female-fertile mutation specifically though there may be a common denominator of actiol influencing quartets of microspores was reported in maize. among many lines, such as abnormal tapetal activity, In soybean, the ms2 and ms6 mutations have a strong there is much work yet to be done at the developmenta~ effect al this stage, causing quartet degeneration. At the molecular and genetic levels before one or several soybean ms3 locus, the mutation causes both quartet and systems can be understood and engineered for widespread microspore degeneration unlike ms2 and ms6. Wall use. formation of young microspores is initiated but not completed. In maize, abnormal development of the micro­ References spore walls is influenced by several genes. Mutations at 1. AulERTSllN MC (1988) Chromosome arm location the ms2 and ms7 loci significantly arrest formation of for ms23 Maize Genet Coop News Lett 62 71. microspore walls. When mutations are at maize mslO and 2. AIBERTSBN MC, SEl.LNER LM (1988) An indepen­ msl3 loci, microspore wall development lags behind dent, EMS-induced dominant male sterile that maps microSp<>re vacuolation. Microspores in sterile anthers similar to Ms41 Maize Geru:t Coop News Lett 62 70. break down al the midvacuolate stage. Both callose 3. AulERTSllN MC, PAIMBR RG (1979) A comparative dissolution and initiation of microspore walls are absent. light and electron-microscope study of microsporo­ genesis in male-sterile (msl) and male-fertile soy­ Microgametogenesis is arrested in ms5 ms5 and msll beans Am J Bot 66 253-265. msll maize anthers. In soybean, however, details are not 4. AulERTSEN MC, PHIWPS RL(1981) Developmental known about postmeiotic milosis in coenocytic micro­ cytology of 13 genetic male sterile loci in maize Can spores in msl msl and ms4 ms4 mutants. Pollen tubes do J Genet Cytol 23 195-208. form and they are able to penetrate the embryo sac in low 5. AulERTSllN MC, NEUFFER MG (1990) Dominant frequencies. male sterile Maize Genet Coop News Lett 64 52. 6. BEADUl GW (1932) Genes in maize for pollen The phenotype of msp mutation in soybean is inconsis­ sterility Genetics 17 413-431. tent. Abnormalities can occur during microsporogenesis 7. BEYERSDORF WD, BINGHAM ET (1977) Male sterility (late sporogenous stage) and microgamelogenesis. This as a source of haploids and polyploids of Glycine suggests that the mutation arises at an early stage of max Can J Geru:t Cytol 19 283-287. development and also that its later expression can be 8. BoBRMA HR, COOPBR RL (1978) Increased female modified by certain faclors. None of the reported male­ fertility associated with the msl locus in soybeans sterile, female-fertile maize mutations has such a broad Crop Sci 18 344-346. spectrum of effects. 9. BRIM CA, YOUNG MF (1971) Inheritance of male­ sterile character in soybeans Crop Sci 11 564-566. There is a limited amount of information on the role of 10. BUNTMAN DJ, HORNER HT JR (1983) Microsporo­ the tapetum in nuclear male-sterile mutations in maize. In genesis of normal and male sterile (ms3) mutant ms3 ms3 soybean, accumulation of sporopollenin in soybean (Glycine max) Scanning Electron Microsc tapetal cells suggests that the tapetum malfunctions early Part II 13-22. in microsporogenesis and disturbs the time of callase 11. Buss GR (1983) Inheritance of a male-sterile mutant activity. This abnormality oftapetal tissue also is associat­ from irradiated 'Essex' soybean Soybean Genet New$ ed with the ms2, ms3, ms6 and msp loci. Two of the three 10 104-108. cytoplasmic male steriles in maize (C-CMS, T-CMS) 12. CAR~ON JB, LERSTEN NR (1987) Reproductive display abnormalities in the tapetal layer. Nevertheless morphology In JR WILCOX (Ed) Soybeans: Improve­ malfunction of the tapetum in anthers of sterile plants ment, production and uses 2nd ed Agronomy 16 95- seems to be a common feature for both nuclear and cyto­ 134. plasmic sterility. No cytological information is available 13. CHAUDHARI HK, DAVIS WH (1977) A new male­ on CMS system in Soybean. sterile strain in Wabash soybeans J Hered 68 266- 267. Nuclear male-sterile mutants affect a variety of cell types 14. CHBN LF, PALMER RG (1985) Cytological studies of in the anther and in some instances such as msl msl triploids and their progeny from male-sterile (msl) soybean, express themselves in a similar manner in MALE STERILITY 17

soybean Theor Appl Genet 71 400-407. 34. GoTTSCHAUC W, KAULMLH (1980b) Asynapsis and 15. CHEN LF, AIBERTSBN MC, PAl.MER RG (1987) desynapsis in Oowering plants II Desynapsis Nucleus Pollen and coenocytic microspore germination in 23 97-120. male-sterile msl soybean Euphytica 36 333-343. 35. GRAYBOSCH RA, BERNARD RL, CREMEENS CR, 16. CHEN LFO, HEER HE, PALMER RG (1985) The PAl.MER RG (1984) Genetic and cytological studies frequency of polyembryonic seedling and polyploids of a male-sterile, female-fertile soybean mutant J from msl soybean Theor Appl Gend 69 271-277. Hered 75 383-388. 17. CHENG PC, GREYSON RI, WALDEN DB (1979) 36. GRAYBOSCH RA, PALMER RG (1985a) Male sterility Comparison of anther development in genie male­ in soybean (Glycine max) I. Phenotypic expression of sterile (msJO) and in male-fertile com (Zea mays) the ms2 mutant Am J Bot 72 1738-1750. from light microscopy and scanning electron micros­ 37. GRAYBOSCH RA, PAIMER RG (l 985b) Male sterility copy Can J Bot 57 578-596. in soybean (Glycine max) II. Phenotypic expression 18. CHOWDHURY JB, VARGHBSE TM (1968) .Pollen of the ms4 mutant Am J Bot 72 1751-1764, sterility in crop plants--A review Palynol Bull 9 71- 38. GRAYBOSCH RA, PALMER RG (1987) Analysis of a 86. male-sterile character in soybeans J Hered 78 66-70. 19. CvrrnR GL (1975) Effect of a genetic male-sterile 39. GRAYBOSCH RA, PAl.MER RG (1988) Male sterility character on the organization and function of the in soybean : An overview Am J Bot 15 144-156. female gametophyte in soybeans PhD Thesis Univer­ 40. HACKE, LINC, YANG H, HORNER HT (1991) The sity of Wisconsin, University Microfilms. Ann Arbor, T-URF13 protein from mitochondria of Texas male­ MI (Diss Abstr 36 4252). sterile maize (Zea mays L) : Its purification and 20. Cl!nllR GL, BINGHAM ET (1977) Effect of soybean submitochondrial localization and immunogold male sterile gene msl on organization and function of labelling in anther tapetum during microsporogenesis the female gametophyte Crop Sci 17 760-764. Plant Physiol 95 861-870. 21. DAVIS WH (1985) Route to hybrid soybean produc­ 41. HANSON MR, CONDE MF (1985) Functioning and tion United States Patent 4,545,146. variation of cytoplasmic genomes : lessons from 22. DElANNAY X, PALMER RG (1982) Genetics and cytoplasmic-nuclear interactions affecting male cytology of the ms4 male-sterile soybean J Hered 73 fertility in plants Int Rev Cytol 94 213-267. 219-223. 42. HANSON MR (1991) Plant mitochondrial mutations 23. DWICK DN (1965) Cytoplasmic pollen sterility in and male sterility Ann Rev Genet 25 461-486. com Adv Genet 13 1-56. 43. HArnELD PM (1982) Comparative physiology of a 24. EMERSON RA (1932) A recessive zygote lethal soybean chlorophyll mutant at two ploidy levels MS resulting in 2:1 ratios for normal vs. male-sterile and Thesis Library Iowa State University Ames IA. colored F 2 colorless pericarp in F 2 of certain maize 44. HBSLOP-HARRISON J (1971) Features of male-sterility hybrids Science ..,5 566. in angiosperms Proc Annu Corn Sorghum Res Con/ 25. EYS"IER WH (193la) Heritable characters of maize 26 14-21. Male sterile 2 J Hered 22 99-102. 45. JAIN SK (1959) Male sterility in Oowering plants 26. EYSIBR WH (193lb) Heritable characters of maize Bibliogr Genet 18 101-166. XXXIX-Male sterile 3 J Hered 22117-119. 46. JOHNS CW, DELANNAY X, PALMER RG (1981) 27. EYSIBR WH (1934) Genetics of Zea Mays Bibliogr Structural sterility controlled by nuclear mutations in Genet 11 187-392. angiosperms Nucleus 24 97-105. 28. FRANKEL R,IZHAR S, NtTSAN J (1969) Timing of 47. KALMAN L, P ARDUCY A, PINTER L (1982) RB callase activity and cytoplasmic male-sterility in cytoplasmic male sterility in maize : Fertility restora­ Petunia Biochem Genet 3 451-455. tion and histology of anther development Maydica 27 29. FORDE BG, OLIVER RJC, LBAVER CJ (1978) Varia­ 1-10. tion in mitochondrial translation products associated 48. KAUL MLH (1987) Male sterility in higher plants with male-sterile cytoplasms in maize Proc Natl Acad Springer-Verlag Berlin. Sci USA 75 3841-3845. 49. KBNNEIL JC, HORNER HT (1985) lnOuence of the 30. GoWBOVSKAYA JN (1979) Genetic control of soybean male-sterile gene (msl) on the development meiosis Int Rev Cytol 58 247-290. of the female gametophyte Can J Genet Cytol 27 31. GoLUBOVSKAYAJN, DISTANOVAEE (1986) locating 200-209. of the mei-gene ms4 by TBA stocks Maize Genet 50. KBNwoRrnY WJ, BRIM CA, WERNSMAN EA (1973) Coop News Lett 60 106-107. Polyembryony in soybeans Crop Sci 13 637-639. 32. GoTTSCHAUC W, KAUL MLH (1974) The genetic 51. KoDURA PRK, RAo MK (1981) Cytogenetics of control of microsporogenesis in higher plants Nucleus synaptic mutants in higher plants Theor Appl Genet 17 133-166. 59 197-214. 33. GoTTSCHAUC W, KAUL MLH (1980a) Asynapsis and 52. LAsER KD, LERSTBN NR (1972) Anatomy and desynapsis in Oowering plants I Asynapsis Nucleus cytology of microsporogenesis in cytoplasmic male­ 23 1-15. sterile angiosperms Bot Rev 38 425-454. 18 RG PALMERET

53. LEE SU, GRACEN VE, EARLE ED (1979) The cytolo­ 68. RHOADES MM (1950) Meiosis in maize J Herd 41'.; gy of pollen abortion in C-cytoplasmic male-sterile 58-67. : com anthers Am J Bot 66 656-667. 69. SADANAGA K, GRINDELAND R (1981) Natural aOll> c, 54. LEE SU, EARLE ED, GRACEN VE (1980) The cytolo­ pollination in diploid and autotetraploid soybew :' gy of pollen abortion in S cytoplasmic male-sterile Crop Sci 13 503-506. ' com anthers Am J Bot 67 237-245. 70. SINGLETON WR, JONES DF (1930) Heritable chll'» 55. NAKASHIMA H, HORNER HT, PALMER RG (1984) ters of maize XXXV Male-sterile J Hered 21 266- Histological features of anthers from normal and ms3 268. mutant soybean Crop Sci 24 735-739. 71. SISCO PH, GRACEN VE, EVERETI HL, EARil! ED, 56. MORTON CM, LAWSON DL, BEDINGER (1989) PRING DR, MCNAY JW, LEVINGS III CS (198S) Morphological study of the maize male-sterile mutant Fertility restoration and mitochondrial nucleic acids ms7 Maydica 34 239-245. distinguish at least five subgroups among CMS-S 57. NEUFFER MG, HOISINGTON DA, BIRD R McK (1987) cytoplasms of maize (Zea mays L) Theor Appl GtMt Designation of new dominant mutants Maize Genet 71 5-15. Coop News Lett 61 50-51. 72. SKORUPSKA H, NAWRACAlA J (1980) Observation of 58. OHMASA M, WATANABE Y, MURATA N (1976) A pollen grains of soybean plants in male-sterile lino biochemical study of cytoplasmic male sterility of Urbana msl Genet Pol 21 33-37. com : Alteration of cytochrome oxidase and malate 73. SKORUPSKA H, PALMER RG (1987) Monosomica dehydrogenase activities during pollen development from synaptic KS mutant Soybean Genet News 14 Jpn J Breed 26 40-50. 174-178. 59. PALMER RG, WINGER CL, ALBERTSEN MC (1978) 74. SKORUPSKA H, PALMER RG (1989) Genetics and Four independent mutations at the msl locus in cytology of the ms6 male-sterile soybean J Hered 118 soybeans Crop Sci 18 727-729. 304-310. 60. PALMER RG, JOHNS CW, MlilR PS (1980) Genetics 75. SKORUPSKA H, PALMER RG (1990) Additional sterile and cytology of the ms3 male-sterile soybean J Hered mutations in soybeans Glycine max (L) Merr J Herd 71343-348. 81296-300. 61. PALMER RG, KIANG YT (1990) Linkage map of 76. STElLY DM, PALMER RG (1980a) A partially male­ soybean (Glycine max L Merr) pp 6.68-6.93 In SJ sterile mutant line of soybeans Glycine max (L) Merr O'BRIEN (ed) Genetic Maps Cold Spring Harbor : Inheritance Euphytica 29 295-303. Laboratory Publisher Cold Spring Harbor New York. 77. STElLY DM, PALMER RG (1980b) A partially malc­ 62. PALMER RG, l