FIELD STUDY OF TEXAS NATIVE EVENING PRIMROSE AND
EVALUATION OF SELECTED SEED TREATMENTS
by
CYNTHIA LOWERY MURPHY, B.S., B.B.A.
A THESIS
IN
HORTICULTURE
Submitted to the Graduate Faculty of Texas Tech University in Partial Fulfillment of the Requirements for the Degree of
MASTER OF SCIENCE
Approved
Accepted
May, 2000 ACKNOWLEDGEMENTS
There are a number of people that I would like to express my gratitude to, for their assistance with this project. Without the open-mindedness of Dr. Dick
Auld, this project probably would not have made it off the ground. I would also
like to thank the other members of my committee Dr. Norman Hopper and
Cynthia McKenney for their contributions.
A special thanks goes to Dr. Andy Herring for his contributions at a
moments notice.
A number of faculty members and professionals were responsible for
assisting with equipment needs Including Dr. Don Wanjura of the USDA, for use of the spectral radiometer. Dr. Henry Nguyen provided the osmometer needed In the priming experiments and Dr. Dan Krieg for his library and as an information
source.
I would like to thank David Becker, Darin Mitchell, and the rest of the farm crew for all their assistance. Life could have been much more difficult without their help especially because they have all the tractors.
My family has been very supportive during this fime,especiall y my in-laws,
"Murph" and Latrece Murphy. My husband, Kendal, became an extra set of hands during planfing, harvest and any time I needed something built that involved his power tools. My son, Benjamin Ray Lowery Parra, has probably suffered the most through my education. My goal has always been to provide him with a better life. Many thanks to my parents for providing Ben with a stable environment to live in.
I would like to thank Landry Lockett and Sandra Balch for taking on some of my day-to-day responsibilities so that I could meet my deadlines. Sandra's ability to Identify nafive sources of evening primrose became a foundation for our
research.
I especially would like to thank my scholarship donors during this program.
The assistance from The Belsterling Foundation of the Dallas Women's Garden
Club, The Herb Society of America - South Texas Unit, and the Native Plant
Society made the great burden of tuition and books not so overwhelming.
Without organizations such as these, higher education would not be possible for many of us.
Finally, a word of thanks to the Texas Department of Agriculture and the
USDA's Fund for Rural America for financialsuppor t of this project. It is my hope that they confinue to findth e Importance in supporting research involving native and medicinal plants. TABLE OF CONTENTS
ACKNOWLEDGEMENTS "'
ABSTRACT v
LIST OF TABLES vl
LIST OF FIGURES vii
CHAPTER
I. INTRODUCTION 1
II. LITERATURE REVIEW 3
Plant Populafion Study 6
Seed Priming Study 7
Red Light Study 8
III. MATERIALS AND METHODS 10
Plant Populafion Study 10
Seed Priming Study 12
Red Light Study 14
IV- RESULTS AND DISCUSSION 18
Plant Population Study 18
Seed Priming Study 30
Red Light Study 34
V. SUMMARY AND CONCLUSIONS 37
LITERATURE CITED 40
APPENDIX 44
iv ABSTRACT
Evening primrose oil has increasingly become of interest to the health
care industry for its legitimate effectiveness in treatment of certain pathological
conditions. The essenfial component of evening primrose oil is gamma-linolenic
acid (GLA) (018:3 A6, 9,12). Four accessions of Texas native evening primrose
containing higher levels of GLA were evaluated for feasibility as a field crop
through transplants. Highest seed yields were reported with plant populations of
32.3 thousand plants per hectare. Very little significance was reported in percent
oil yield between the three different plant spacings examined. Germination
chamber and greenhouse experiments were conducted using KNO3 and PEG
8000 as priming agents at -0.5 and -1.0 MPa for 5-, 10-, and 15-day treatments
to overcome dormancy symptoms affecting germination. The most effective
treatments for O. elata were achieved with -0.5 MPa osmotic potenfials using
PEG 8000 for 5 days and -1.0 MPa using PEG 8000 for 5 or 10 days. The best
priming treatment for O. yames//resulted from using PEG 8000 at the -0.5 MPa
level for 15 days. The most effective treatment for O. rhombipetala occurred at
the 10 and 15 day treatments at the -1.0 MPa osmotic potential. Red light
treatments showed no significant differences for the species O. rhombipetala. O.
elata showed the t)est results using a germination chamber regardless of the light treatment. LIST OF TABLES
4.1 Seed yield of four accessions of Oenothera at three plant populafions at Lubbock, Texas in 1999. 23
4.2 Oil content of four accessions of Oenothera grown at Lubbock, Texas in 1999 as determined by nuclear magnetic resonance. 25
4.3 Yield of highest yielding plants of four accessions of Oenothera grown at Lubbock, Texas in 1999. 26
4.4 Total oil yield of highest yielding samples submitted for analysis of three species of evening primrose. 28
4.5 Percent germination in a germination chamber after 14 days for seed of three evening primrose species treated in priming solufions for 5-, 10-, and 15-days. 32
4.6 Percent emergence in the greenhouse after 21 days from seed of three evening primrose species treated in four priming solutions for 5-, 10-, and 15-days. 33
4.7 Percent germination of three evening primrose species under six light treatments. 36
A1 Soil and water analysis by A & L Plains Agricultural Laboratories, Inc. 45
A2 Accessions of the four highest seed yielding plants within each 3.5 m unit and their respective oil percentages. 46
VI LIST OF FIGURES
3.1 Spectral radiometer analysis of various light sources and filters to isolate the optimal 660 nm wavelengths. "• ^
4.1 Seed yield of three species of evening primrose grown at three different plant populations in Lubbock, Texas, 1999. 24
4.2 Percent oil of four evening primrose accessions grown at three plant populations at Lubbock, Texas in 1999. Analysis provided by the University of Idaho, Moscow, Idaho. 27
4.3 Oil yield in kg ha'^ of four accessions of Oenothera species grown in Lubbock, Texas, 1999. 29
VII CHAPTER I
INTRODUCTION
Historically, evening primrose (Oenothera spp.) has been grown both as an attractive wildfiower and an herbal supplement. Evening primrose oil has increasingly been recognized by the medical community as a legitimate health care product. Medical researchers have found that the high levels of gamma- linolenic acid (GLA) (018:3 A6, 9, 12) found in evening primrose oil can be used to treat the many pathological condifions in humans caused by deficiencies in
GLA. Stress, diabetes, consumption of alcohol and aging are reported to inhibit the producfion of GLA (Favafi, King, and Mazzanti 1991). Dietary supplementation with GLA can reduce lymphocyte proliferation in older humans which often have decreased A-6 desaturase activity (Wu and Meydani 1996).
Studies have shown that long-term use of evening primrose oil has allowed rheumatoid arthritis patients to reduce their use of steroidal and non-steroidal anti-infiammatory drugs. GLA may also provide relief from itching and antihistamine use associated with eczema and other skin disorders. Gamma- linolenic acid has also been effective at lowering blood pressure levels and in the treatment of hypertension. Recent research indicates that GLA may also reduce the symptoms of diabetes mellitus and atherosclerosis (hardening of the arteries)
(Hathaway 1999; Keen et al. 1993).
Many of these pathological conditions have k>een associated with defiencies of n-6 essenfial fatty acids which occurs In individuals with limited 1 delta-6 (A-6) desaturase enzyme. This enzyme is responsible for the formation of long-chain polyunsaturated fatty acids (PUFAs) from linoleic acid (LA 018:2
A6,9). Supplementation of the diet with the GLA derived from the oil of plants such as evening primrose is thought to be an answer to this rate limited reaction and reduces the severity of many diseases (Mengeaud, Nano, Fournel and
Rampa! 1992).
As a plant, native to West Texas region, Oenotheras have the ability to thrive in our challenging climate and produce economic quantifies of oil and GLA.
Three species in particular, Oenothera elata (A. Gray ex S. Watson), Oenothera jamesii (Torrey & A. Gray), and Oenothera rhombipetala (Nuttall ex Torrey & A.
Gray) have been identified by researchers at Texas Tech for their higher levels of
GLA; therefore, have become the focus of continued study. There is a need for cultural producfion information and to improve the germinafion of evening primrose seed if commercial producfion of this medicinal crop is to occur in the
West Texas region. The purposes of this study were to determine the cultural requirements and attempt to reduce the seed dormancy characteristics of evening primrose. CHAPTER II
LITERATURE REVIEW
There are several sources of gamma-linolenic acid (GLA) in plants. The higher plant families Onagraceae, Boraginaceae, Saxifragaceae, and
Scrophulariaceae all contain species producing GLA. Many nafive accessions of
Evening Primrose have been collected and evaluated for their desirable oil by researchers at Texas Tech University (Yaniv, Auld, Heuer and McKenney 1999).
Species which produce commercially competitive quantifies of GLA have been identified as Oenothera rhombipetala, Oenothera elata, and Oenothera jamesii.
All three of these species are within the Oenothera section of the Onagraceae family which contains 70 species (Dietrich, Wagner and Raven 1997). This section is divided into 5 subsections listed as Raimannia (11 species), Emersonia
(4 species), Oenothera (13 species), Munzia (39 species), and Nutanfigemma (3 species). O. rhombipetala is classified as part of the Raimannia subsection while
O. jamesii and O. elata are classified within the Oenothera subsection. The
Oenothera secfion Oenothera are indigenous only to North America and occur in open areas of arid regions (Dietrich, Wagner, and Raven 1997). They reproduce only sexually and bear more seed than other members of Oenothera. All three species have yellow fiowers which open near sunset; an indeterminate growth habit; appear in a rosette stage when young; and have a haploid chromosome number of n=7. O. rhombipetala is reported to be self-incompatible (Dietrich and
Wagner 1987; 1988). O. jamesii and O. elata are self compatible but also outcross (Dietrich, Wagner, and Raven 1997). O. jamesii is suggested to have evolved from O. elata from possible changes in moth pollinators (Dietrich,
Wagner, and Raven 1997). One of the latter's most noticeable differences is the length of the hypanthium (where the corolla and calyx attach to the stamens In a funnel appearance) and seed size. O. jamesii has a much longer hypanthium and slightly larger seed size than O. elata (the seeds of both species are angular). Since O. rhombipetala is classified within an entirely different subsection, its physical differences are much more obvious. Its fiowersappea r in dense crowns at the end of stems and are much smaller as are the seeds (which appear smooth and oblong) and seed pods. Under irrigation, O. rhombipetala blooms much eariier in the season than either O. jamesii or O. elata. Its branches are also less rigidtha n O. jamesii ar\6 O. elata whose appearance is more of a pyramidal shape while O. rhombipetala starts out pyramidal but ends up with twisfing branches. The seeds in all three species are borne in capsules and are two-ranked (two rows In each locule) (Oorrell, and Johnston 1979).
Evening primrose seed is made up of approximately 15% protein, 43% cellulose and llgnin, and 24% oil (Phillips, and Huang 1996). The retail market for this oil is approximated at $0.48 (U.S.) per gram (Favati, King, and Mazzanti
1991). This oil contains from 7-14% GLA. The high fiber proportion of the seed is believed to be primarily seed coat (Hudson 1984). Seed maturation in evening primrose has been shown to affect the levels of GLA present. During early stages of seed development, linoleic acid is In greatest abundance but as development progresses, the A-6 desaturase enzyme becomes more active and GLA accumulates (Phillips 1996). According to a study conducted in Israel by
Zohara Yaniv, higher linoleic and GLA levels were achieved with fall planfings of
Oenothera lamarckiana as opposed to spring plantings (Yaniv, Ranen, Levy and
Palevitch 1989). With increases in temperature, there was a decrease in most seed yield components including seed weight, number of seed per inflorescence, and number of pods. Observafions of Oenothera rhombipetala during late July and August in Lubbock, Texas seem to support this. Further studies in Israel concluded that 30-40 days following anthesis when the stigma may receive shed pollen were the most responsive to temperature. Their program has involved producing lines whose seed will mature in cool weather (Levy, Palevitch, and
Ranen 1993). Oenothera elata and Oenothera jamesii both mature during
October and Novemt)er at cooler temperatures for the Lubbock, Texas area.
The different studies for this thesis have t)een examined due to the limitations of small seed size and the survival mechanism of dormancy. Seed dormancy can be either physical or physiological (Oopeland and McDonald
1995). Dormancy has been classified in different manners but for this thesis It is understood to be of two types: primary and secondary (Oopeland and McDonald
1995). Primary dormancy is also divided into two categories: exogenous and endogenous. Exogenous dormancy pertains to tiie restrictions Induced by the seed coat. Because seeds require moisture, temperature, oxygen, and in many cases light, to gemilnate; an impermeable seed coat may inhibit this.
Endogenous dormancy is caused by the presence of a chemical inhibitor within the seed and is strongly affected by seed maturity. Secondary dormancy is described as the condifion when all requirements for germination have been met but one. Comparative seed coat anatomy describes the section Oenothera subsection Oenothera and subsection Raimannia to have a thick endotesta, persistent mesotesta and a nonspeciallzed exotesta with a few exceptions to those examined within the study (Tobe, Wagner, and Chin 1987).
Plant Populafion Study
Published Information on current production of evening primrose oil involves transplanfing young plants of Oenothera biennis from the greenhouse to the field In what can be an expensive and labor intensive task (Brandle, Oourt, and Roy 1993; NIghfingale and Baker 1995). Very little published information as to the specifics of this process and evening primrose is available. This would appear to be tiie best method to achieve a stand as the seed of evening primrose exhibits dormancy characterisfics. In Ontario, two methods of production are used; transplanfing greenhouse plants to the fieldi n the spring and direct seeding In the fall (Roy, et al. 1994). Plug transplanting by mechanical means is a common method for many flower and vegetable crops. Mechanical transplanting would keep labor costs low and bypass the risks of seed germination in the field. Holland Transplanters (Holland, Ml) estimates that a
Rotary One transplanter is capable of planting sixty transplants per minute with one operator (Holland Transplanters, Inc. 1998). Planting depth and spacing can be selected for a uniform rate, the fun-ow opened, a plug dropped in, packed and watered with the same implement. Seed Primino Study
"Osmotic priming is a technique of imbibing seeds in an osmotic solufion under controlled conditions to maintain seeds in the desiccation-tolerant lag phase of germination" (Wartidinlngsih, Geneve, and Kester 1994, p. 1445).
Osmotic priming has been useful in many crops to Improve germinafion (Yoon,
Lang, and Cobb 1997; Bradford, Steiner, and Trawatha 1990; Samfield, Zajicek, and Cobb 1991). Charles Danvin conducted experiments in 1855 that speculated at osmotic seed priming when he immersed seeds in salt water. The outcome was to suggest that seeds could be distributed across the seas. Of importance to this study Is that not only did they germinate aftenA/ards but also some showed Improved germination rates (Hartmann, Kester, Davles, and
Geneve 1997). Seeds can be hydrated in specific solutions such as polyethylene glycol (PEG), potassium hydroxide, potassium nitrate, potassium phosphate or sodium hypochlorite for several days at concentrations that are not toxic to the seeds but also don't allow the radicle to emerge (Oopeland and McDonald 1995).
PEG and mannitol have been used when trying to avoid salt toxicity associated with solutions of NaOl, KNO3, etc. Frequenfiy used osmotic potentials range between -0.5 MPa and -1.5 MPa and treatments can last up to 15 days
(Hartmann, Kester, Davies, and Geneve 1997). PEG with a molecular weight of
4000 and higher is more commonly used as those with lower molecular weights can be taken up by germinating seeds and cause damage (Oopeland and
McDonald 1995). High molecular weight PEG has reported toxicity due to the manufacture of PEG itself and the presence of aluminum and magnesium ions in forming the polymer of ethylene glycol (Mexal, Fisher, Osteryoung, and Reid
1975). Ethylene glycol by Itself is commonly known as anti-freeze (Hart, Hart, and Oralne 1995). The osmotic potential of finesolutio n can be verified in the laboratory by use of an osmometer. Seeds are then dried completely and can be tested for rapid, uniform emergence in a variety of media from moist paper towels to potting soil. Past experience has shown evening primrose to germinate at various rates according to the species and environment. In the greenhouse, germination can take from one to three weeks, in the field even longer including the following season forming a persistent seed bank (Baskin and Baskin 1993).
Red Lioht Studv
"Seed germination is one of the many aspects of plant growth and development which can be modified by light." (Thomas 1992, p. 239). It has already been observed in pilot studies at Texas Tech that Oenotheras germinated In light far better than in dark and AOSA standard tesfing for similar species calls for tesfing to take place with light (AOSA 1986; 1983). Extensive research has been conducted on species to support the photoblasfic nature of
Oenothera seed (Ensminger, and Ikuma 1988, 1987a, 1987b). Light intensity and spectral quality have been known to affect many plant species In both beneficial and detrimental aspects. Studies have shown that Oenothera biennis
L could not adequately germinate under plant canopies (Gross 1985). The far- red wavelengths (720 - 800 nm) dominate under a plant canopy. In full sunlight,
8 the red wavelengths (580 - 720 nm) dominate over the far-red wavelengths but
penetrate less deeply into the soil. As evening primrose prefer light to germinate
and are sown near the surface, it is possible that red wavelengths or the intensity
of red wavelengths are beneficial to their germination. Red light can be produced
artificially with plant growth lights, which emit light In the blue as well as the red
region (Preece and Read 1993).
The chemical responsible for the light reaction in seeds Is known as
phytochrome, a protein pigment. When most concentrated, phytochrome
appears blue and Is paled by red light but darkened by far red light (Oopeland
and McDonald 1995). This process is constantly changing by periods of light and
darkness. By exposing intensified phytochrome to red light and paling it to a far-
red form, which is the active form for enzyme synthesis, many plant species can
be influenced to germinate. Phytochrome is known to be associated with cell
membranes and is present in the seed as the phytochrome-red form. By exposing It to red light and intensifying it to the phytochrome-far-red form, studies
have shown that it may be affecting membrane permeability or causing gibberellin synthesis to occur (Oopeland and McDonald 1995). The literature
reviewed for this thesis did not insist on a precise locafion of the phytochrome pigment but did agree to it being a cause of a seed dormancy condition in photosensitive seeds (Thomas 1992). CHAPTER IN
MATERIALS AND METHODS
Plant Populafion Studv
Accessions of O. rhombipetala ^-36 and R-46) and O. elata (R-50) and
O. jamesii (T-60) were seeded In the greenhouse in 22.86 x 12.7 x 6.35 cm aluminum baking fins containing Ball Growing On® mix on February 16, 1999.
The tins were placed randomly on benches in the University Greenhouse and misted twice daily to provide moisture for the shallow sown seeds. When seedlings reached at least 1.27 cm in height and had 2 sets of true leaves they were transplanted into round, plastic 5.56 cm wide x 5.87 cm deep plug trays
(Johnny's Selected Seeds, Albion, ME). Water and fertility were maintained consistently throughout the process with all accessions receiving weekly batch mixed fertility applicafions of 20-20-20 (N-P-K) fertilizer solution. At nine weeks after transplanting, all plug trays were moved outside and placed underneath cold frames for a two week hardening off process. On May 7, 1999 (11-1^ weeks after seeding - WAS), the plugs were watered, pulled from the plastic trays and packed into heavy plastic fiatsaccordin g to the randomized complete block design. The plugs were then transplanted using a Rotary One (Holland
Transplanters, Holland, Ml) on 30, 60, and 90 cm plant spacing in rows spaced one meter apart. This provided final plant populations of 32,310 plants, 16,129 plants, and 10,759 plants per hectare. Plugs were watered-in by the transplanter and subsequent Irrigation was accomplished through buried drip tape placed 30
10 cm below the soil surface that delivered water at 1.4 l/h with emitters 45 cm apart. Irrigation was started immediately to reduce transplant shock. Weather station data were collected by a Campbell Scientific MetDatal weather station located approximately 400 m from the test site. Flea beetles were managed by applications of Liquid Sevin carbaryl insecticide at a rate of 19.72 ml/gal.
Applications were made with a Solo backpack sprayer and the final application was made on June 28, 1999 (19 WAS) to avoid killing any insects beneficial for pollination. After this fime, the flea beetle population did not reach critical levels and treatments were not necessary. At maturity, adjacent plants within a 3.5 m sampling unit of the 9.14 m blocks were harvested by hand, threshed and the seed screened with 12.7 cm perforated plate soil sieves (Humboldt Mfg.,
Norridge, II). Seed samples were then cleaned using a General Seed Blower and finalsee d weight determined. The experiment was evaluated as a randomized complete block design with four replications. Each of the four accessions was evaluated in separate experiments. Data were analyzed by analysis of variance (SAS, Version 7 1998) and means separated with a Fisher's
Protected Least Significant Difference Test at the 0.05 level of probability. Oil content was determined by nuclear magnetic resonance using a canola seed sample as a standard.
11 Seed Primino Studv
Seed priming treatments consisted of thirteen different treatments including a nontreated control in a completely randomized design. Polyethylene glycol (PEG) 8000, formerly 6000 (Sigma), and potassium nitrate KNO3 (Sigma) was prepared at two different osmotic potentials for 5-, 10-, and 15-day treatments respectively. Nontreated dry seeds were used as a control. Evening primrose seeds primed in distilled water showed excessive radicle emergence in eariier observafions and made an ineffecfive control. Once the radicle had emerged, seeds did not survive the drying process post priming. Each of the priming solutions was adjusted to osmotic potentials of -0.5 MPa and -1.0 MPa as determined by a vapor pressure osmometer (Wescor, Logan, UT).
Determining the concentrafion of PEG 8000, the solution needed to achieve the
Inifial -0.5 MPa (-5 bars) and -1.0 MPa (-10 bars) was provided by literature
(Michel, and Kaufmann 1973) and verified by vapor pressure osmometry. To achieve -0.5 MPa at 25°0, 202 g of PEG per kg of deionized water was ufillzed and 296 g of PEG for -1.0 MPa. To determine the necessary proportions of
KNO3 necessary to achieve -0.5 MPa and -1.0 MPa a set of reference solutions was built in 200 ml volumes and the van't Hoff factor used to estimate the osmolality desired (Zhang [c. 1999?]; Chang 1988). This factor Is as follows:
Ys = -iCRT,
^s = osmotic potential,
i = a constant that accounts for dissociafion of the solute,
0 = the osmolality determined by the osmometer, 12 R = the gas constant (0.0831),
T = absolute temperature (273.15 + 25°0) for room temperature.
The osmometer is first calibrated to a set of standard NaOl solutions provided by the manufacturer, which must be done at the beginning of the session. Three readings were taken of each standard and were similar in outcome before continuing. O was the desired osmometer reading and was approximately equal to 202 mmol/kg for ^s at -5 bars and 404 mmol/kg for -10 bars. Because the osmometer readings are very sensitive, three readings were taken of the sample to verify accuracy to the desired osmotic potential. Seed samples of 0.25 g each were obtained using accessions of O. elata (R-50), O. jamesii (T-60) and O. rhombipetala (R-46) for 5-, 10-, and 15-day intervals respectively. These seed samples were enclosed In 10 cm x 10 cm muslin squares to avoid loss of the small seeds, fied with nylon cord and labeled. These seeds inside the muslin bags were then placed in Ehrienmeyer flasks containing 300 ml of each of the osmotic treatment solutions. Solufions were maintained at room temperature and aerated with tubing and a small air pump (Hartmann, Kester, Davies, and
Geneve 1997). LIghfing consisted of Sylvania standard grow lights on an eight hour day. At 5-, 10-, and 15-day intervals, samples were removed from the treatment solutions, rinsedwit h deionized water, and dried for 24 hours under a fan at 25° 0. Random samples of 50 seeds were replicated three times for each treatment and placed in petri dishes on the top of blue blotter circles (Hoffman
Mfg., Inc. Albany, OR). These dishes were placed in a germination chamber
(Stults Mfg.) at alternating temperatures of 20° 0 for 16 hours and 30° 0 for 8 13 hours. The eight hours at 30° 0 was also provided light by Sylvania standard grow lights. The 20/30° alternating temperature, with light and on top of blotter circles is the testing procedure recommended by the Association of Official Seed
Analysts (AOSA 1986; 1983) for the similar species O. biennis L. Germination was achieved with radicle emergence of 0.5 cm or greater as observed on a dally
basis for 14 days. The experiment was statistically evaluated as a completely
randomized design with three repetitions using analysis of variance (SAS,
Version 7 1998). Means were separated with Fisher's Protected Least
Significant Difference Test at the 0.05 level of probability.
Fifty seeds, replicated four times, from each of the previous priming treatments were sown into aluminum baking finswit h drainage containing Ball
Growing On® soilless potting media and placed on a greenhouse bench. A
Hobo® tidbit was placed in the middle of the bench to monitor greenhouse temperature. Pans were misted up to twice daily with reverse osmosis water.
Emergence was recorded on day 1, 3, 6, 9, 12, 18, and 21.
Red Lioht Studv
The objective of this experiment was to determine If seed germination could be improved by omitting all light wavelengths except those at the 660 nm
range. A light source emitting wavelengths including those In the 660 nm range was determined, using a spectral radiometer (Analytical Spectral Devices,
Boulder, 00) placed approximately 10.16 cm from the light. Once a light source
14 was found (Sylvania standard 34 watt Grow Light), all other wavelengths had to be filteredou t with the exception of those at 660 nm. This was accomplished using a double layer of a commercial brand of red cellophane (Olearphane,
Highland, IL) and verified with the spectral radiometer (Fig. 3.1). All extraneous light was blocked by enclosing the entire experiment in a constructed plywood box with a 60.96 cm light fixturecontaine d Inside. This isolation box was partitioned into two sections with a plywood divider and enclosed by a hinged front lid. The same grow lights were suspended over the entire experiment with the red cellophane tacked over one section of the box to be directly over petri dishes plated with fifty randomly selected seeds from each of the three accessions. This was replicated three times for each accession. The seeds were placed on top of blue blotter circles, moistened with distilled water and enclosed in the petri dishes. All dishes received an eight hour light period and were misted with the same distilled water as needed. Simultaneously, fifty randomly selected seeds, replicated 6 times, for each of the three accessions, were plated in plastic petri dishes for comparison against a germination chamber environment. All seeds were placed on top of blue blotter circles that had been previously moistened with distilled water and the lids placed on top but not sealed. Three of the replicafions of each species were randomly placed on the same tray in the germinafion chamber. The remaining three replications were grouped without respect to species, in groups of three and enclosed in a double layer of red Olearphane cellophane. The ends and sides of this wrapping were fastened to keep extraneous light from entering while the treatment was in
15 process. The wrapped dishes were placed on the same tray but a tray away from the nontreated replicafions in the same germination chamber. The chamber was set on 20/30° 0 with an 8-hour light cycle for the warm period. The lights were the same Sylvania standard grow lights used in other experiments that
were able to produce light In the 660 nm range. Germination was recorded daily
over a period of 14 days for radicle emergence of 0.5 cm.
16 35000
30000
250001 Coo! While Fluorescent Grow Light 20000 Warm White Fluorescent Grow Light & Red Glass Reflectance Grow Light and Red Olearphane
15000 +
10000
5000
350400450500550600650700750800850900950 1000
Wa\€length [nm]
Figure 3.1. Spectral radiometer analysis of various light sources and filters to isolate the optimal 660 nm wavelengths.
17 CHAPTER IV
RESULTS AND DISCUSSION
Plant Population Studv
Three different plant spacings of transplanted evening primrose were evaluated for seed yield. Even though individual plant yields varied and often improved with increased distance between the plants, the improvement was not enough to offset the greater numbers of seed obtained with decreased spacing and increased plant populafions per hectare (Table 4.1, Fig. 4.1). The spacings were large enough that even with the smallest one selected, the crowding was not enough to prevent normal growth. All of the transplants that survived the initial adjustment to the field environment were able to branch, bloom and set seed for analysis. Mean seed yields and statistical significance at the 0.05 level of probability are located In Table 4.1.
Development in the fieldfo r each of the four accessions (R-50, T-60, R-
36, R-46) varied a great deal due to species differences. R-36 and R-46 varied the least developmentally being the same species. R-36 and R-46 began branching from their rosette stage on June 1, 1999 (15 weeks after seeding -
WAS) with floral inifiation noted on June 22 (17 WAS) at an approximate height of 40.64 cm. T-60 began sporadic branching on June 8 (16 WAS) and did not begin fioral inifiafion until August 2,1999 (24 WAS) at which fime their height approximated 66-91 cm. R-50 had the same branching date as T-60 but began an inconsistent floweringpattern , noted on September 6, within the study and
18 bud abscission without appearances of drying. Even though the plants looked
very healthy and were without wilting signs, Irrigation was increased and within
four days blooming commenced throughout the study.
Harvesting took place for R-36 and R-46 on September 1, 1999 (28 WAS).
The approximate dimensions for these two accessions at harvest were 76 cm.
wide X 83.5 cm. tall. Both of these accessions developed into an open pyramidal
shape but as the plants matured, the stalks began a twisting pattern. A couple of
factors influenced harvesting the two O. rhombipetala accessions though still
blooming. Many birds were attracted to the smaller less woody capsules and
seed. The proximity of larger, second year specimens made a preferred habitat
for dove and many smaller birds. More than one nest was found upon harvest of
these larger specimens. It was also observed that even though O.
rhombipetala's growth habit Is Indeterminate; during the month of August, blank
spaces existed In the stalks where flowers had been but no seed capsules had formed. This possibly could be a response to the heat usually experienced for
Lubbock at this fime of year. The likelihood of this seems strong; as this period
passed, capsules began forming leaving a gap of approximately 2.54 cm. The
lower capsules were maturing and starting to open and it was decided to harvest.
It was also observed that even though the plant appeared healthy and bloomed
profusely, the capsules appeared small as did the seeds in comparison to wild samples collected. This observafion was supported by consistenfiy small yields throughout the O. rhombipetala samples (Table A2). Because the soil and water analyses (Table Al) showed high salinity, low fertility and a number of other
19 problems with this site, it is possible fiiat this species has less tolerance to poor
environmental conditions than O. elata and O. jamesii.
Even though a large amount of leafy, green foliage existed on O. jamesii
(T-60), harvest took place on November 18, 1999 (35 WAS). The presence of
harvesting (T-60) wifii green foliage meant extra care was taken to find a warm,
dry environment with ventilafion to prevent fungal growth from occurring. The
harvested plants had to be fully dry before threshing could take place. The seed
had reached its dark brown ripe color and the capsules were beginning to quickly
split even while retaining green color. This entire plant including the seed
capsules was much larger and woodier than any of the other oilseed evening
primrose in the study. Approximate dimensions for this plant at harvest were
124.46 cm. wide x 127 cm. tall. In appearance, it developed a very open,
pyramidal shape and looked very crowded on the smallest spacing. Even though the capsules from this species were larger than R-50, the seed weight yields were less and this potentially could have been impacted by the rapid splitting of the woodier capsules and/or excessive competition within the row.
Harvest for O. elata (R-50) took place November 30, 1999 (37 WAS); the week after temperatures remained below freezing for a few days. This accession
had been checked daily for ripenessan d the onset of a cold spell turned the
immature, pink seed color quickly to the ripe, dark brown. An added plus to the cold weather was the senescence of foliage and capsules. The slighfiy smaller capsules (but similar in shape to O. jamesii) did not go through the quick splitting.
R-50 also did not have the extremely coarse stalks and stems that made
20 harvesfing the more rigidT-60 , by hand, difficult. Approximate dimensions for this accession at harvest, were 96.52 cm. wide x 106.68 cm. tall.
Seed yields by weight were statistically compared within each species for significance at the 0.05 level. Approximate weights per 100 seeds for O. elata
(R-50) were 0.03 gm; for O. jamesii {1-60), 0.06 gm and for O. rhombipetala (R-
36) and (R-46) were 0.02 gm. Table 4.1 summarizes the plant population results and shows that 32.3 thousand plants ha'\ and 16.1 thousand plants ha'^ plant spacing has the highest yield in kg ha'^ for O. elata (R-50) without significant difference between them. 10.8 thousand plants ha'^ has a significanfiy poorer yield with O. elata than the other two. O. jamesii (1-60) showed significantly higher yields with 32.3 thousand plants ha"^ while 16.1 thousand plants ha"^ and
10.8 thousand plants ha"^ were significantly poorer than 32.3 thousand plants ha'
^ but not when compared to each other. O. rhombipetala (R-46) showed no significant differences between any of the spacing treatments while O. rhombipetala (R-36) showed significant improvement when planted with the narrowest spacing but showed no significant differences between the other two.
This is also illustrated graphically In Fig. 4.1. The projected yield ha'^ was assuming rows of one meter apart.
Per cent oil content was provided using Nuclear Magnefic Resonance by the University of Idaho using a canola standard and is summarized in Table 4.2.
The percent oil was determined from the four highest seed yielding individual samples of each 3.7 m replication. Oil yield for O. elata improved when planted on the narrowest spacing but was not significantly so from the widest spacing. It
21 cannot be concluded with this species and in this environment that populafion
significanfiy affects oil yield. This conclusion Is further supported by the lack of
significant differences between plant spacing and oil percentages within O. jamesii {T-eO) or O. rhombipetala (R-46). O. rhombipetala (R-36) showed
highest (and identical) oil percentages when planted on 0.6096 m or 0.9144 m.
Mathematically illustrating total yield when combining both the components of
seed yield and per cent oil per hectare are shown in Fig. 4.3. Both R-50 and T-
60 support the cultural pracfice of planting higher plant populations to achieve an
optimum total yield ha"^ even though individual plants may comparatively have
lower yields than those on an increased spacing. Where there is no significant
difference in yields between spacing, a producer would choose the lesser plant
population unless the cost of transplants was more than a little offset by the
increase in oil yield.
Statistical analysis of the four highest yielding plants from each replication for seed and oil yield in kg ha'^ is outlined in Tables 4.2, 4.3, and 4.4. This
illustrates what a producer could achieve through a selection program with higher yielding accessions.
22 Table 4.1. Seed yield of four accessions of Oenothera at three plant populations at Lubbock, Texas in 1999.
SPECIES
Plant Populafion 0. elata 0. jamesii 0. rhomb. O. rhomb. (R-50) (T-60) (R-46) (R-36)
(1 X 10^ Plants ha"''^ •
32.3 1184 a^ 1035 a^ 67 a^ 69 a^
16.1 1068 a 672 b 60 a 45 b
10.8 674 b 444 b 46 a 33 b
Coefficient of 10.0% 26.0% 34.6% 28.0% Variation
"•"Means within a column not followed by the same letter differ at the 0.05 level of probability by Fisher's Protected Least Significant Difference Test.
23 1400 r
1200
1000
LSD (0.05)
800 •—O. elata (R-50) Seed Yield •— O. jamesii (T-60) (kg/ha) k-0.rt)omb.(R-46) 600 K— O. rtiomb. (R36)
400
200 -
10.8 16.1 32.3
Thousand Plants per Hectare
Seed Yield
Figure 4.1. Seed yield of three species of evening primrose grown at three different plant populafions In Lubbock, Texas, 1999.
24 Table 4.2. Oil content of four accessions of Oenothera grown at Lubbock, Texas in 1999 as determined by nuclear magnetic resonance.
SPECIES
Plant Population O. elata O. jamesii O. rhomb. O. rhomb. (R-50) (T-60) (R-46) (R-36)
(1 X 10^ Plants ha"*) Yield (%)
32.3 11.7a^ 11.3a^ 10.0 a^ 9.6 b^
16.1 11.0b 11.0a 10.1a 10.2a
10.8 11.4ab 11.1a 10.0a 10.2a
Coefficient of 6.1% 5.2% 9.0% 6.8% Variation
"•^Means within a column not followed by the same letter differ at the 0.05 level of probability by Fisher's Protected Least Significant Difference Test.
25 Table 4.3. Yield of highest yielding plants of four accessions of Oenothera grown at Lubbock,Texas in 1999.
SPECIES
Plant Population 0. elata 0. jamesii i0. rhomb. 0. rhomb. (R-50) (T-60) (R-46) (R-36)
(1 X 10^ Plants ha""")- -—Yield (kg ha""")-
32.3 1590 a^ 1643 a ^ 124 a ^ 116a^
16.1 1131 b 801 b 75 ab 55 b
10.8 673 c 444 b 46 b 33 b
Coefficient of 7.1% 21.8% 37.2% 39.9% Variation
"•"Means within a column not followed by the same letter differ at the 0.05 level of probability by Fisher's Protected Least Significant Difference Test.
26 14 r
12
10 LSD (0.05) - O. elata (R-50) 8 - O. jamesii (T-60) Percent Oil -O. rhomb. (R-46) - O. rhomb. (R-36)
4 -
2 -
0 10.8 16.1 32.3 Thousand Plants per Hectare
Seed Oil Percentage
Figure 4.2. Percent oil of four evening primrose accessions grown at three plant populations at Lubbock, Texas in 1999. Analysis provided by the University of Idaho, Moscow, Idaho. 27 Table 4.4. Total oil yield of highest yielding samples submitted for analysis of three species of evening primrose.
SPECIES
Plant Population O. elata O. jamesii O. rhomb. O. rhomb. (R-50) (T-60) (R-46) (R-36)
(1x10^ Plants ha'"") Total Oil Yield (kg ha"^)
32.3 186 a^ 185 a^ 13 a^ 11a^
16.1 124 b 88 b Sab 6 b
10.8 77 c 49 b 5 b 3 b
Coefficient of 8.4% 23.0% 39.9% 37.0% Variation
"^Means within a column not followed by the same letter differ at the 0.05 level of probability by Fisher's Protected Least Significant Difference Test.
28 LSD (0.05) O. elata (R-50) O. jamesii (T-60) Oil Yield O. rhomb. (R-46) (kg/ha) O. rhomb. (R-36)
10.8 16.1 32.3 Thousand Plants per Hectare
Total Oil Yield
Figure 4.3. Oil yield in kg ha"^ of four accessions of Oenothera species grown In Lubbock, Texas, 1999.
29 Seed Primino Studv
Osmotic priming using a variety of solutions Is a common treatment to promote seed germination. However, each species has its requirements as to the most effective solution, osmotic potential, and duration of the treatment to inifiate germinafion.
Results from the germinafion chamber showed germination to have commenced by the 3^^ and 4^ day for the treated seeds but not unfil the 5*^ day for the nontreated control. The majority of germination for the treated seeds was complete in 4 to 5 days. The majority of germination for the controls was complete within 5 days. According to the statistical analysis, the study showed that for O. elata (R-50), all of the treatments gave significant germination
Increases above the nontreated seeds with the exception of the -0.5 MPa KNO3.
Seeds in this solufion developed a white fungus after several days In the petri dishes for the 10- and 15-day treatments even though this fungus did not appear as they were removed from the muslin squares. Germination for -1.0 MPa PEG
5- and 10- day treatments as well as KNO3 5- and 15- day and -0.5 MPa PEG 5- day showed no significant differences between the treatments and gave the highest germination. The longer the treatment, the more viscous the PEG solutions became making aeration more difficult. At 15 days, some stringiness was noted on the outside of the muslin bags, which could have been fungal growth from an anaerobic condition. The alternative to maintaining a constant level of solufion with adding distilled water as needed would be to use fresh
30 solutions dally. It was observed that If a fill line was not kept, the osmotic potential changed dramatically by water evaporafion and left an Increase in solutes within the solution. Compared to the nontreated control there was significant improvement within every species due to at least one priming treatment. Even though treatments using the same solutions could be for a 5-,
10-, or 15-day duration, they were analyzed as separate treatments; each with a lot of 0.25 gms of seed from which random samples were removed for plating in petri dishes.
For the greenhouse portion of this experiment, emergence was used for data collection. Results were slower than expected but explained by the results of the Hobo®. Temperatures ranged from 14°0 to 34°0 on a dally basis. For this reason, data was taken for 21 days, Instead of 14 as In all of the other experiments. If the majority of emergence had not taken place by this time, the treatment could not be called an improvement. Table 4.7 shows the results. For
O. elata and O. jamesii, the mean separation between treatment effects was not clearcut. The most effecfive treatment for O. elata stood out at 88% emergence.
The least effective treatments were at the -0.5 MPa KNO3 treatments. This osmotic potential was not enough to prevent frequent radicle emergence while being primed. Unexpectedly, the most effecfive treatment for O. yames//was at the -0.5 MPa level for the 5-day PEG treatment.
31 Table 4.5. Percent germinafion in a germination chamber after 14 days for seed three evening primrose species treated in priming solufions for 5-, 10- and 15-days.
SPECIES
Priming Treatment 0. elata 0. jamesii 0. rhombipetala (Days) (R-50) (T-60) (R-46)
-0.5 MPa KNO3 (5) 35 ef^ 48 c^ 78 abe^ -1.0MPaKNO3(5) 83 ab 53 c 83 ab -0.5 MPa PEG (5) 87 a 65 ab 78abc -1.0 MPa PEG (5) 89 a 55 be 81 ab
-0.5MPaKNO3(10) 5g 30 d 66 cd -1.0MPaKNO3(10) 56 d 59abc 85 a -0.5 MPa PEG (10) 61 cd 54 be 72 be -1.0 MPa PEG (10) 91 a 57 be 82 ab
-0.5MPaKNO3(15) 23 f 36 d 45 e -1.0MPaKNO3(15) 82 ab 56 be 85 a -0.5 MPa PEG (15) 73 be 70 a 58 d -1.0 MPa PEG (15) 85 ab 59abc 75 abe
Nontreated Control 41 e 36 d 72 be
Coefficient of Variation 12.8% 13.6% 10.1%
"•^Means within a column not followed by the same letter differ at the 0.05 level of probability by Fisher's Protected Least Significant Difference Test.
32 Table 4.6. Percent emergence in the greenhouse after 21 days from seed of three evening primrose species treated in four priming solufions for 5-, 10-, and 15-days.
SPECIES
Priming Treatment 0. elata 0. jamesii 0. rhombipetala (Days) (R-50) (T-60) (R-46)
-0.5 MPa KNO3 (5) 41 e^ 46 cd^ 61 b^ -1.0MPaKNO3(5) 88 a 56 ab 67 ab -0.5 MPa PEG (5) 77 cd 67 a 63 ab -1.0 MPa PEG (5) 85 abe 53 bed 59 b
-0.5MPaKNO3(10) 14f 33 e 57 b -1.0MPaKNO3(10) 78 bed 48e 92 a -0.5 MPa PEG (10) 73 d 47 bed 55 b -1.0 MPa PEG (10) 87 ab 50 bed 57 b
-0.5MPaKNO3(15) 22f 34e 36 c -1.0MPaKNO3(15) 84abc 43 de 59 b -0.5 MPa PEG (15) 76 cd 57 abe 38 c -1.0 MPa PEG (15) 85 abe 44 de 66 ab
Nontreated Control 79 abed 51 bed 57 b
Coefficient of Variation 9.9% 16.6% 15.6%
^Means within a column not followed by the same letter differ at the 0.05 level of probability by Fisher's Protected Least Significant Difference Test.
33 Red Light Studv
The purpose of this procedure was to determine the germination response
of the three evening primrose species used throughout this study when exposed
to a constant source of red light. Results are in Table 4.8. O. elata (R-50)
showed the highest germinafion, nontreated, In the germinafion chamber under
fluorescent lighting. This response was, however, not significantly different at the
0.05 level from the red light treatment in the same chamber or nontreated in the
germination chamber equipped with grow lights. The red light treatment In the
germination chamber with grow lights showed significanfiy poorer germination
response than the first three but was not significantly different from the isolation
box red treatment. The nontreated isolation box showed the poorest response to
germination. O. jamesii (T-QO) also showed the best germinafion in the
germination chamber with fluorescent lighting and nontreated. It was not
significantly different from the red light treatment in the germination chamber with
either fluorescentlight s or grow lights. The poorest response was in the Isolation
box under both treatments which suggests that the constant humidity and/or the
intensity of the light source has a strong impact on this species. The fluorescent
light according to the spectral radiometer emitted a higher intensity of light than the grow light source but did not quite emit at the 660 nm range. Another Item to consider is that because counts were taken daily and removed from their treatments some of the response to the red light treatments could have been
reversed. O. rhombipetala (R-46) showed the best response with a red light
34 treatment in the germinafion chamber with grow lights. However, it was not significanfiy different at the 0.05 level than any of the other treatments. Because the best responses took place in the germinafion chamber regardless of fluorescent or grow lights, It is suggested that evening primrose is more sensitive to moisture at germination than light quality for all three of these species.
Because the petri dishes were not sealed in any of the environments and required daily misting in the isolafion box, there could have been enough moisture stress to affect germinafion as well as the interruption in the light treatment.
35 Table 4.7. Percent germination of three evening primrose species under six light treatments.
SPECIES
Light Treatment O. elata O. jamesii O. rhombipetala (R-50) (T-60) (R-46)
•% Germination-
Germ, chamber with fluor. 89 a^ 66 a^ 73 a^ Lighting - nontreated
Germ, chamber - red light 88 a 60 ab 77 a Treatment
Isolation box - nontreated 65 c 17c 75 a
Isolation box - red light treated 69 be 16c 73 a
Germ, chamber with grow 87 a 50 b 72 a lights - nontreated
Germ, chamber with grow 77 b 54 ab 80 a lights - red light treatment
Coefficient of Variation 5.2% 19.4% 9.5%
"^Means within a column not followed by the same letter differ at the 0.05 level of probability by Fisher's Protected Least Significant Difference Test.
36 CHAPTER V
SUMMARY AND CONCLUSIONS
This thesis has tried to gain an understanding Into the processes of
dormancy and ufilization of undomesticated evening primrose seed for oil
production purposes. Field producfion Is possible using transplants for this area.
A means of mechanically harvesting and large scale seed cleaning are a
necessity for commercialization to occur and needs to be addressed. The hand
harvesting and seed cleaning techniques used in this study show expected yields
under a research environment. Mechanically harvesting typically loses a certain
amount of seed but has the potential to be offset by an improved field
environment. Mechanically transplanfing involves an inifial investment of about
$3000 for the transplanter, plus the expense of greenhouse grown transplants. A
transplanted field would opfimize growing space and prevent cosfiy overseeding.
In the event of a severe winter, a seeded fieldstand s the chance of young plants
being frozen. Treafing evening primrose culturally as an annual could relieve
some risk of an additional six to eight months of maintenance in the field.
Evening primrose seed is very small and difficult to work with but produces a
large plant requiring room to branch within one season. Plant spacings selected
in this study were within suggested plant spacings used by producers in Canada
(Baker 1997). O. rhombipetala yields were so much lower than O. elata or O. jamesii that production of this species for the oil itself does not seem profitable.
37 The difficulty in producing hybrid crosses with species of Oenothera exhibiting self -incompatibility is documented (Cleland 1972). The possibility of low yields due to the lack of compatible pollen should also be considered and has been documented in other species of Oenotheras (Silander 1978). When anthesis began, the only other species in proximity to the study had not begun to bloom and is verified In photographs. Soil fertility could also be examined as a means for increasing yields. Another negative factor concerning this species is that the seed matures during warmer temperatures which has an effect on yield components. Positive features of R-50 and T-60 were their later maturing habit under cultivafion. Planting studies at different sites using the same cultural pracfices might show how much the environment has an impact on the yield components of these species.
Priming treatments were shown to be effective In increasing the germination of these species of evening primrose seed. The less than optimal temperatures in the greenhouse during the greenhouse emergence phase of the priming study are likely to have caused the slower emergence and less than expected emergence results. In order to prevent fungal growth from occurring within the priming solutions, changing the solufions daily, even though inconvenient to store in large amounts, would be an option to maintaining consistent osmotic potentials by adding distilled water to a specific level. Further research should be done to determine how long primed evening primrose seed remains viable after treatment.
38 The technique used for the red light study did not show the expected results. Interrupfion in the treatment by daily data recording with exposure to an outside light source could have affected germination rates. The intensity of the light source is a strong factor to consider. Superior results obtained in the germination chamber suggest that light quality by itself Is not the strongest factor affecting evening primrose germination.
This thesis has attempted to cover as much preliminary ground as possible in the effort towards production of a Texas native medicinal plant.
Because the evening primrose species evaluated; evolved In this environment, they have already developed a hardiness that some crops can never achieve for this area. It is hoped that continued research will be able to develop the assets growing around us everyday.
39 LITERATURE CITED
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43 APPENDIX
44 Table Al. Soil and water analysis by A & L Plains Agricultural Laboratories, Inc.
Oateoorv Julv. 1997 March. 1999 Guidelines*
Soil: Organic Matter % 0.7 1.7* Low Phosphorous (ppm) 16 38* Very High Potassium (ppm) 386 350* Very High Magnesium (ppm) 530 212* Very High Calcium (ppm) 1200 980* Low Sodium (ppm) N/A 520* Very High Soil pH 7.2 7.6 Nitrate (ppm) 7 8* Low Sulfur (ppm) 4* N/A Low Zinc (ppm) 0.6* N/A Very Low Manganese (ppm) 6* N/A Low Iron (ppm) 4* N/A Low Copper (ppm) 0.5* N/A Low Boron (ppm) 2.3* N/A High
Julv. 1997 Februarv, 2000 Irrioation Water: Sodium (ppm) 170 250 > 180 Severe Pre Calcium (ppm) 57 128 100-200 Probler Magnesium (ppm) 164 102 > 60 Severe Prol CO3 (ppm) <1 <1 HOO3 (ppm) 268 360 > 180 Severe Pro Chloride (ppm) 550 480 > 300 Severe Pro Water pH 7.4 7.2 Normal 6.5 - 8.^ Phosphorous (ppm) 0.044 0.121 0.1 -0.4 Norma Potassium (ppm) 20 20 5-20 Normal Nitrate NO3 (ppm) 23 25 5-50 Normal Sulfate S04(ppm) 705 470 > 180 Very Higf Boron (ppm) 0.88 0.70 0.75 - 2.0 Problei
*ltems followed by an asterisk are explained in the guidelines column.
45 Table A2. Accessions of the four highest seed yielding plants within each 3.5 m unit and their respective oil percentages
ID No. Accession Plant Seed Percent Oil No. Weioht (cms)
59 0E11 1 49.1 11.1 60 0E11 7 42.7 12.1 85 0E11 9 53.9 11.6 7 0E11 11 40.0 11.9 110 0E12 2 47.8 12.0 31 0E12 4 56.7 12.1 137 0E12 8 49.0 11.3 61 0E12 12 55.5 12.3 115 0E13 2 46.7 11.2 158 0E13 7 53.3 11.3 36 0E13 10 48.1 11.9 179 0E13 11 39.8 10.9 138 0E14 1 53.7 11.5 190 0E14 2 56.5 11.6 86 0E14 4 50.9 12.3 139 0E14 11 43.4 11.9 87 0E21 2 64.0 7.4 62 0E21 3 52.4 11.4 191 0E21 5 59.5 11.2 180 0E21 6 62.2 10.8 89 OE22 1 104.6 11.8 192 OF?? 3 75.7 11.2 88 OE22 4 72.7 11.0 35 OE22 6 74.1 10.8 140 OE23 1 68.9 11.4 113 OE23 2 64.8 11.9 143 OE23 3 59.5 11.4 181 OE23 5 68.8 10.3 114 OE24 1 79.5 10.7 182 OE24 CV I 66.3 11.3 63 OE24 3 72.7 11.4 111 OE24 5 76.2 11.4 32 OE31 1 62.1 11.5 142 0E31 2 56.7 10.6 159 0E31 3 47.5 11.9
46 Table A2. Oonfinued.
ID No. Accession Plant Seed Percent Oil No. Weioht (cms)
141 0E31 4 76.3 11.2 34 OE32 1 58.1 11.5 160 OE32 2 58.0 11.4 90 OE32 3 59.8 12.4 64 OE32 4 60.7 11.3 161 OE33 1 45.1 11.4 8 OE33 2 61.5 11.5 33 OE33 3 48.3 10.9 112 OE33 4 75.4 10.5 91 OE34 1 73.8 11.6 9 OE34 2 95.1 11.5 93 OE34 3 66.5 11.9 92 OE34 4 56.4 11.5 67 0J11 1 66.3 10.9 55 0J11 3 49.9 10.6 121 0J11 4 51.9 11.4 134 0J11 12 55.2 11.2 29 OJ12 4 63.6 12.3 97 0J12 10 46.9 11.9 156 0J12 11 65.2 11.7 81 0J12 12 48.7 10.5 39 0J13 3 37.0 11.9 120 0J13 5 88.7 12.0 165 0J13 6 48.2 10.2 177 0J13 12 46.4 10.7 56 0J14 2 40.6 11.8 147 0J14 3 39.1 11.2 6 0J14 5 30.9 10.9 96 0J14 12 34.6 11.2 135 0J21 2 34.4 10.5 82 0J21 3 28.1 10.9 164 0J21 4 33.4 10.1 157 0J21 5 44.4 11.1 57 OJ22 1 46.3 10.4 11 OJ22 2 36.5 10.9 178 OJ22 5 77.5 11.1 119 OJ22 6 58.7 11.5
47 Table A2. Oonfinued.
ID No. Accession Plant Seed Percent Oil No. Weioht foms)
146 OJ23 1 71.8 11.4 189 OJ23 3 70.5 10.6 38 OJ23 5 71.0 10.9 136 OJ23 6 59.1 11.4 66 OJ24 1 55.7 11.2 95 OJ24 2 33.5 11.5 117 OJ24 3 20.4 10.9 184 OJ24 6 53.7 11.1 118 0J31 1 65.2 11.8 163 0J31 2 67.7 10.5 58 0J31 3 46.1 11.4 37 0J31 4 63.8 12.3 83 0J32 1 39.6 10.9 3 OJ32 2 20.5 12.5 183 OJ32 3 45.3 10.9 10 OJ32 4 26.3 11.1 162 OJ33 1 50.9 10.5 65 OJ33 2 31.7 10.9 145 OJ33 3 30.7 10.0 144 OJ33 4 60.3 10.8 30 OJ34 1 23.3 10.6 116 OJ34 2 23.7 11.8 94 OJ34 3 26.3 10.7 84 OJ34 4 38.8 10.7 49 OR(R-36)11 1 3.2 9.8 106 OR(R-36)11 3 2.8 9.4 42 OR(R-36)11 6 2.7 9.9 155 OR(R-36)11 10 2.7 9.5 18 OR(R-36)12 1 7.7 8.9 13 OR(R-36)12 5 4.5 9.4 125 OR(R-36)12 7 5.3 9.4 1 OR(R-36)12 10 4.8 10.0 71 OR(R-36)13 6 2.9 9.9 168 OR(R-36)13 9 2.5 10.3 28 OR(R-36)13 11 3.5 10.3 188 OR(R-36)13 12 3.7 9.9 22 OR(R-36)14 1 3.5 9.9
48 Table A2. Oonfinued.
ID No. Accession Plant Seed Percent Oil No. Weioht (oms)
186 OR(R-36)14 2 2.6 9.4 129 OR(R-36)14 7 2.4 9.9 50 OR(R-36)14 9 2.5 8.1 185 OR(R-36)21 1 3.3 10.0 21 OR(R-36)21 2 4.0 11.1 167 OR(R-36)21 4 3.6 10.6 70 OR(R-36)21 6 4.1 10.3 79 OR(R-36)22 3 3.1 10.4 51 OR(R-36)22 4 3.4 10.2 41 OR(R-36)22 5 3.4 10.8 130 OR(R-36)22 6 3.1 10.0 166 OR(R-36)23 2 2.7 9.3 20 OR(R-36)23 3 4.5 10.7 107 OR(R-36)23 5 4.1 9.8 123 OR(R-36)23 6 1.9 10.2 124 OR(R-36)24 1 4.7 8.7 52 OR(R-36)24 2 2.5 10.1 99 OR(R-36)24 4 3.7 10.0 80 OR(R-36)24 5 2.3 10.8 132 OR(R-36)31 1 1.4 10.5 131 OR(R-36)31 2 3.4 10.7 12 OR(R-36)31 3 4.1 9.9 69 OR(R-36)31 4 1.5 10.6 53 OR(R-36)32 1 2.9 9.8 19 OR(R-36)32 2 1.4 11.1 40 OR(R-36)32 3 4.0 10.6 148 OR(R-36)32 4 1.8 10.9 108 OR(R-36)33 1 1.1 10.8 133 OR(R-36)33 2 5.0 9.4 2 OR(R-36)33 3 6.0 9.7 98 OR(R-36)33 4 3.8 10.5 54 OR(R-36)34 1 4.0 10.9 109 OR(R-36)34 2 3.4 7.8 68 OR(R-36)34 3 2.5 10.0 122 OR(R-36)34 4 3.5 9.8 25 OR(R-46)11 1 1.5 10.3 24 OR(R-46)11 4 1.7 10.4
49 Table A2. Oonfinued.
ID No. Accession Plant Seed Percent Oil No. Weioht foms)
45 OR(R-46)11 11 1.4 9.1 75 OR(R-46)11 12 2.9 8.9 102 OR(R-46)12 1 4.4 9.7 15 OR(R-46)12 9 7.3 10.3 103 OR(R-46)12 11 6.1 9.4 74 OR(R-46)12 12 5.8 10.4 26 OR(R-46)13 2 3.1 10.1 46 OR(R-46)13 8 4.5 8.1 127 OR(R-46)13 11 2.6 10.3 152 OR(R-46)13 12 2.2 10.2 16 OR(R-46)14 5 4.5 10.5 171 OR(R-46)14 8 4.8 11.2 47 OR(R-46)14 9 5.2 10.6 128 OR(R-46)14 10 3.2 10.6 172 OR(R-46)21 1 1.3 9.6 17 OR(R-46)21 2 5.0 9.6 151 OR(R-46)21 3 1.2 9.7 170 OR(R-46)21 5 2.1 10.5 153 OR(R-46)22 1 4.9 10.5 173 OR(R-46)22 2 5.1 8.5 44 OR(R-46)22 3 5.8 10.4 174 OR(R-46)22 6 11.2 9.7 14 OR(R-46)23 1 3.5 9.2 150 OR(R-46)23 2 3.7 10.1 23 OR(R-46)23 5 4.6 12.2 76 OR(R-46)23 6 15.0 10.0 175 OR(R-46)24 1 1.8 11.4 176 OR(R-46)24 2 3.2 9.6 4 OR(R-46)24 4 3.5 10.8 101 OR(R-46)24 6 2.6 9.8 187 OR(R-46)31 1 3.3 9.9 154 OR(R-46)31 2 3.0 10.1 48 OR(R-46)31 3 2.5 7.3 104 OR(R-46)31 4 2.6 10.9 77 OR(R-46)32 1 5.5 8.5 43 OR(R-46)32 2 11.6 9.7 73 OR(R-46)32 3 4.5 10.2
50 Table A2. Oonfinued.
ID No. Accession Plant Seed Percent Oil No. Weioht (oms)
100 OR(R-46)32 4 3.1 10.8 169 OR(R-46)33 1 2.6 9.5 27 OR(R-46)33 2 5.3 10.3 5 OR(R-46)33 3 2.2 11.3 105 OR(R-46)33 4 2.0 10.6 149 OR(R-46)34 1 3.8 10.1 126 OR(R-46)34 2 4.8 10.4 78 OR(R-46)34 3 6.8 9.3 72 OR(R-46)34 4 4.9 10.3
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