Production of interspecific hybrids between Glycine max and G. tomentella through embryo culture

Summary Interspecific hybrids have been obtained in an incompatible cross between Glycine max and G. tomentella through the in vitro culture of hybrid embryos. The percentage of successful pod setting in the crosses averaged 12.8% but there were marked differences depending on the soybean cultivar used as the female parent. Hybrid embryos at globular to heart stages were extracted from the embryo sac 15–25 days after pollination and cultured in vitro. Hybrid plants were successfully obtained by culturing the embryos on B5 medium supplemented with 0.1 mg/l IBA followed by culture on B5 medium supplemented with 0.1 mg/l TBA plus 0.25 mg/l 2-iP. The F1 plants resembled the wild male parent in growth form, but had an intermediate leaf shape between that of the parents.     

Interspecific hybridization in the genus Gossypium through embryo culture

Summary With embryo rescue and culture techniques interspecific hybrids have been obtained amongst diploid cultivated (Gossypium arboreum, G. herbaceum) and wild species (G. stocksii, G. anomalum) of cotton. The early abscission of the young bolls was prevented by repeated application of growth regulators followed by the culture of immature hybrid embryos (15 days after pollination). The best growth and development were obtained on modified Murashige and Skoog's medium with supplements. The plants were stouter when the cultures were first incubated in the dark for 15–25 days, followed by exposure to light. The hybrid plants transferred to soil developed further and matured, and were more or less intermediate between their respective parents.     

Nutrient culture medium components affecting plant recovery from immature embryos of three Glycine genotypes and an interspecific hybrid grown in vitro

Summary Immature embryos of Glycine max (L.) Merr. cv. Bay, G. tabacina (Labill.) Benth. and G. tomentella Hayata were cultured on 72 media combinations to identify a nutrient medium which would allow a greater percentage of interspecific plants to be recovered from cultured embryos of G. max × various perennial Glycine species. The highest mean plant recovery rate of 79% was from a medium containing B5 nutrient salts as reported by Gamborg et al. (1968), vitamin components according to Williams (1978) and 30 g/l sucrose. This is as compared with 25% from a medium used previously. In an additional test, 67% of hybrid embryos of G. max × G. tomentella were recovered from the same medium, from which G. tomentella was most effectively recovered in all testing.Contribution No. 89-276-J of the Kansas Agric. Exp. Stn., Manhattan.     

Interspecific hybridization in the genus Arachis through embryo culture

Summary The embryos of a cultivated tetraploid peanut (Arachis hypogaea), a wild diploid species (A. villosa), and their hybrid embryos, which generally abort in nature, were cultured in vitro and the plants have been successfully transferred to the soil. The hybrids showed triploid chromosome number (3x=30). The significance of wide hybridization in peanut-improvement programs is discussed.     

Interspecific hybridisation betweenLimonium perigrinum Bergius andLimonium purpuratum L.

Summary Interspecific hybrids betweenLimonium perigrinum andL. purpuratum were obtained usingL. perigrinum as the female parent. No hybrids were produced by the reciprocal cross. Twelve- to 15-day-old embryos were rescued and cultured within their embryo sacs on modified B5 or KM medium. After two to three days the embryos were excised from their embryo sacs and re-plated on to fresh medium. When the embryo-derived plantlets had attained a length of 1 cm they were transferred to a modified MS medium containing BA and NAA for shoot proliferation. Plantlets were transferred to modified MS medium supplemented with IBA for 24 hours for root initiation then to a modified growth-regulator-free MS medium for root growth. After a further 28 days the plantlets were transferred to soil-less medium for acclimatisation. The hybrid characteristics of one of the 15 embryo-derived plants were determined by flow cytometry and by examination of morphological features. The mean DNA contents of 2C nuclei fromL. perigrinum, the hybrid andL. purpuratum were 13.98 pg, 16.81 pg and 19.37 pg, respectively. Mitotic and meiotic chromosome counts fromL. perigrinum andL. purpuratum showed that both parents and their hybrids had identical chromosome numbers (2n=24), and that the species were closely related. Morphological analyses of leaves and flowers showed that the hybrids displayed a number of features intermediate between both parents.Abbreviations B5 Gamborg et al. (1968) B5 medium - BA benzyl adenine - IBA indole-3-butyric acid - KM Kao and Michayluk (1975) medium - MS Murashige and Skoog (1962) medium - NAA napthalene-1-acetic acid - PVP polyvinylpyrrolidone     

An intersubgeneric hybrid between Glycine tomentella Hayata and the soybean,G. max (L.) Merr.

Summary The objective of the present paper is to provide information on the morphology and cytology of an intersub-generic hybrid (2n=59) between Glycine tomentella Hayata (2n=78) and G. max (L.) Merr. (2n=40) obtained through in vitro immature seed culture. The hybrid plant was slow in vegetative growth and twinning like the female parent but morphologically was intermediate between both parents for several traits. At metaphase I, the average chromosome associations and ranges for 25 cells were 44.0 I (37–51)+7.5 II (3–11). The plant was completely pollen and seed sterile. The present investigation suggests that wild perennial Glycine species can be exploited as either the male or female parent in wide hybridization programs with the soybean, G. max.     

Interspecific hybridization between sunflower and wild perennial Helianthus species via embryo rescue

Interspecific crosses were made between the cultivated sunflower inbred line HA89 and accessions of five wild perennial Helianthus species (H. giganteus L., H. laevigatus T. & G., H. resinosus Small, H. pauciflorus Nutt. and H. decapetalus L.) resistant to broomrape ( Orobanche cernua Loefl.). Using the genetic male-sterile isogenic version of that line as female, successful reciprocal crosses were also obtained with all these wild species except with H. decapetalus. Five-day-old hybrid embryos were excised and cultured in vitro. In all cases, few mature plants were obtained from embryos in early developmental stages (early heart and globular) but up to 28% mature plants were obtained from embryos in later stages of development. Very immature embryos were difficult to excise without damage. Hybrid embryos and mature plants were obtained from all five wild species. Interspecific hybrid embryos from different wild species showed distinct developmental potentials, the proportion of hybrid embryos in different developmental stages varying among species. Differences in the proportion of hybrid embryos at the several developmental stages were also obtained for the reciprocal crosses (cultivated × wild), which showed higher proportion of fully developed embryos. Hybrids involving H. giganteus and cultivated sunflower were difficult to obtain without the use of embryo culture. Embryo culture proved to be an useful tool to overcome post-zygotic hybrid incompatibility in different Helianthus spp., and facilitated interspecific transfer of resistance to O. cernua. This revised version was published online in July 2006 with corrections to the Cover Date.     

Effect of maturity on competitive ability in soybeans

Summary The effect of maturity on competitive ability in two cultivars of soybeans (Glycine max (L.) Merr.) was evaluated. Maturity dates within a genotype were simulated by removing half the plants from pure stands at end of flowering and twice later at approximately weekly intervals. Simulation of the early maturity of an equal competitor gave a significant competitive advantage to the late genotype only 2 out of 15 times. The average effect from simulation of earlier maturity within the genotype was statistically significant, but small, especially since the assumed difference in physiological maturity was 27 days. Most genotypic difference in competitive ability probably are due to characteristics other than maturity.Approved by the Director as a contribution from the Missouri Agricultural Experiment Station (Journal Series No. 7877).     

Karyotype analysis and interspecific hybridisation in three perennial Sesbania species (Leguminosae)

Summary The somatic chromosome number in Sesbania sesban var. nubica, S. goetzei and S. keniensis (Leguminosae; Papilionoidae) was found to be 2n=12. These findings were in agreement with earlier reports on S. sesban and S. keniensis. The chromosome number 2n=12 is a new record for S. goetzei. Similarities in karyotypes were found in the three species. All species had one pair of long metacentric chromosomes; the second pair was submedian, followed by four smaller pairs of metacentric chromosomes. Nucleolar organiser regions in the form of satellites were found on the short arm of the fourth chromosome pair in S. sesban and S. keniensis. Interspecific crosses in all possible combinations were carried out, resulting in pod and viable seed formation for the crosses S. sesban x S. goetzei, S. sesban x S. keniensis, S. goetzei x S. sesban and S. goetzei x S. keniensis. The two crosses with S. keniensis as a female parent were unsuccessful. The hybrid plants established normally and produced viable seeds.     

A partially male-sterile mutant line of soybeans,Glycine max (L.) Merr.: inheritance

Summary A partial male sterility system in the soybean (Glycine max (L.) Merr.) germplasm population AP6(SI)CI was found to be controlled monogenically by a recessive allele, msp. Observations of msp msp plants in different environments suggested that environmental conditions significantly affect expressivity of the msp allele with respect to male sterility. We obtained no experimental evidence of cytoplasmic effects on msp expression. Background genotypes, however, seem to affect msp expressivity through their determination of flowering dates and resultant interactions with varying environmental conditions.Homogeneous populations of partially male-sterile plants can be generated by increasing families of msp msp plants in fertility-inducing environments, if measures are implemented to prevent the introduction and/or build-up of fertile genotypes that arise from natural cross-pollinations.Joint contribution: North Central Region, Agricultural Research, Science and Education Administration, U.S. Department of Agriculture, and Journal Paper No. J-9596 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa 50011; Project 2107.     

Interspecific hybridization of Brassica napus and B. juncea through ovary,ovule and embryo culture

Summary Employing in vitro culture of ovaries, ovules and embryos, interspecific hybrids have been obtained amongst two important oilseed crops, Brassica napus x B. juncea and their reciprocal. The test-tube hybrid plants have been transferred to the field, and reared to maturity. The F₁ seeds obtained from the hybrid ovaries showed normal germination, and the hybrid plants exhibited a range of variation of characters.     

A partially male-sterile mutant line of soybeans,Glycine max (L.) Merr.: Characterization of the msp phenotype variation

Summary Investigations of variable expression of msp partial male sterility in soybeans (Glycine max (L.) Merr.) showed that higher temperatures promote male fertility in msp homozygotes and showed that infectious agents are not intrinsic to the sterility system. Exchange grafts failed to modify fertility levels of msp msp rootstocks, Msp Msp scions, and their self-progeny. Tests for soybean mosaic virus and tobacco ringspot virus were negative in partially male-sterile plants, in control fertile plants, and in self-progeny of grafted plants. Growth-chamber experiments and field observations manifested that male fertility of msp msp plants is higher in hot environments than in cooler ones. The unexpected aberrant ratios of fertile to partially male-sterile plants observed in 1977 (Stelly & Palmer, 1980) are explainable on the basis of msp temperature sensitivity.Our observations suggest that homogeneous msp msp populations may be increased in hot environments.Research Geneticist, SEA-AR, USDA, Iowa State University, Ames, Iowa 50011, USA.     

Interspecific hybrids of Cyclamen persicum Mill. and C. purpurascens Mill. produced by ovule culture

Summary Interspecific crosses were made to introduce the scent of flowers of C. purpurascens into C. persicum cultivars and ovule culture was used to rescue the abortive hybrid embryos. Cultivars of C. persicum diploid (CPD, 2n=2×=48) and C. persicum tetraploid (CPT, 2n=4×=96) were the pistillate parents and wild species of C. purpurascens (CP, 2n=34) were staminate parents. After pollination, crossed ovaries were collected periodically and examined using paraffin sections. Histological observations suggested that both hybrid ovules of CPD x CP and CPT x CP should be transferred to culture medium 35 days after pollination. Based up on this observation, crossed ovaries were collected 28 days after pollination and ovules with placenta were transferred to MS (1962) medium containing 3% sucrose. These ovules were cultured in the dark at 25° C. The hybrids (2n=41) derived from CPD x CP had the scent of C. purpurascens, whereas the hybrids (2n=65) derived from CPT x CP had the scent of C. persicum. Although both hybrids had complete genomes from the parents and produced a few viable pollen grains, they failed to yield viable seeds by self- and cross-pollination with fertile pollen grains of C. persicum cultivars.Abbreviations CPD C. persicum diploid - CPT C. persicum tetraploid - CP C. purpurascens     

Use of ovule culture to cross between Delphinium species of different ploidy

The applicability of ovule culture was examined in reciprocal crosses between the diploid species D. grandiflorum L. and the tetraploid cultivar D. hybridum ‘Galahad’. An interspecific hybrid plant of D. hybridum ‘Galahad’ × D. grandiflorum was obtained,and hybridity was confirmed by chromosome counting. The hybrid was different from the parents in flower color and size but intermediate between the parents in leaf shape. The hybrid showed low pollen fertility and failed to produce viable seeds by either self- or backcross-pollination using fertile pollen grains from its parents. The applicability and efficiency of the ovule culture technique for genetic improvement of the genus Delphinium through interspecific hybridization between different ploidies is discussed. This revised version was published online in July 2006 with corrections to the Cover Date.     

Each species of Glycine collected in Taiwan has a unique seed protein pattern

Several annual and perennial species in the genus Glycine Willd., including G. soja, long-pod G. tomentella, short-pod G. tomentella and G. tabacina, collected in Taiwan and nearby islands were studied for variations of their seed proteins. SDS-PAGE and Western blotting were used to analyze the total proteins, the heat soluble proteins, six seed maturation proteins (GmPMs) and one seed storage protein. The various species had different patterns of seed heat soluble proteins. In addition,each species of Glycine collected in Taiwan exhibited unique seed maturation protein patterns. They had several cross-reactive polypeptides recognized by specific antibodies against GmPM1, GmPM2 and GmPM8, but only one polypeptide recognized by antibodies against GmPM4, GmPM5 and MP130. The long pod G. tomentella, which has been suggested as a new species and renamed as G. dolichocarpa, could be distinct from the short pod G. tomentella on the basis of the analysis of these biochemical markers. It is also indicated that these GmPM antibodies may be used to distinguish between and within other Glycine species. This revised version was published online in July 2006 with corrections to the Cover Date.     

Establishment of Chinese soybean Glycine max core collections with agronomic traits and SSR markers

It is very important to efficiently study and use genetic diversity resources in crop breeding and sustainable agriculture. In this study, different sampling methods and sample sizes were compared in order to optimize the strategies for building a rationally sized core collection of Chinese soybean (Glycine max). The diversity in the core collection captured more than 70% of that in the pre-core collection, no matter what sampling methods were used, at a sampling proportion of 1%. Core collections established with both simple sequence repeat (SSR) marker data and agronomic traits were more representative than those chosen on an independent basis. An optimal sampling method for a soybean core collection was determined, in which strategy ‘S’ (allocating accessions to clusters according to the proportion of square root of the original sample size within each ecotype) was used based on SSR and agronomic data. Curve estimation was used to estimate the allelic richness of the entire Chinese soybean germplasm and a minimum sample size for a core collection, on which a sampling proportion of about 2% was determined to be optimal for a core collection. Further analysis on the core collection with fourteen agronomic traits and allelic constitution at 60 SSR loci suggested that it highly represented the entire collections both on genetic structure and diversity distribution. This core collection would provide an effective platform in proper exploitation of soybean germplasm resources for the study of complex traits and discovering important novel traits for crop genetic development.     

Interspecific hybridization inBrassica juncea × Brassica hirta using embryo rescue

Summary Interspecific hybrids have been obtained in an otherwise incompatible cross betweenBrassica juncea × Brassica hirta through the in vitro culture of hybrid ovules and ovaries. The best response was observed from ovules and ovaries cultured 10–15 and 5–7 days after pollination respectively on a basal medium supplemented with indoleacetic acid, kinetin and casein hydrolysate. In some cases the basal cut end of the ovaries proliferated to form callus and shoots. The in vitro-derived hybrid seeds varied in their colour, size and shape, and the F₁ plants in the field showed a large diversity in their morphological traits. The hybrids were sterile, and had an intermediate number of chromosomes (2n=30).     

Pollen production in soybeans with respect to genotype,environment, and stamen position

Summary Our objective was to determine the average numbers of pollen grains from fertile plants (Ms ₁) and the average numbers of coenocytic microspores from genetic male sterile plants (ms ₁ ms ₁) in soybeans, Glycine max L. Merr. Comparisons were made between the average numbers of pollen grains and the average numbers of coenocytic microspores with respect to environment where plants were grown and to stamen position in the flower. Five male sterile lines were used. They included the North Carolina ms ₁ mutant, the cultivar Hark with the ms ₁ gene, and mutants identified as the Urbana, Tonica, and Ames male steriles. Three environments used were the Agronomy Farm, University of Georgia, Athens, Georgia; the Agronomy and Agricultural Engineering Research Center, Iowa State University, Ames, Iowa; and the Agronomy Greenhouse, Iowa State University, Ames, Iowa.Pollen production from fertile plants varied from 374 to 760 pollen grains per anther among genetic lines and environments. This variation may be an important consideration in selecting a male parent to use as a pollinator for hybrid seed production.Among fertile plants, the average numbers of pollen grains per anther of the separate stamen and and of the lower whorl of stamens were significantly different only in greenhouse-grown plants. Among male sterile plants, the average numbers of coenocytic microspores per anther of the separate stamen and of the lower whorl of stamens were significantly different in three genotype x environment combinations. These three exceptions did not conform to any genetic or environmental pattern. Deviations from the expected ratio of 4 pollen grains from fertile plants: 1 coenocytic microspore from sterile plants were attributed to initial differences in the average number of microspore mother cells between the two genotypes.Joint contribution: Agricultural Research Service, USDA, North Central Region, and Journal Paper No. J-8910 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa 50011; Project 2107.     

Wide crosses in soybeans: Prospects and limitations

Summary It was attempted, though in vain, to cross soybean with Glycine species of the subgenus Glycine.     

Interspecific hybrids of Cyclamen persicum and C. graecum

Summary Interspecific crosses were made to introduce the disease resistance of Cyclamen graecum into C. persicum cultivars and the abortive hybrid embryos were rescued by ovule culture. Diploid and tetraploid cultivars of C. persicum (CPD, 2n=2x=48; CPT, 2n=4x=96) were the pistillate parents and wild form of C. graecum (CG, 2n=84) were the staminate parents. After pollination, crossed ovaries were periodically collected and examined using paraffin sections. Histological observations suggested that both hybrid ovules of CPD × CG and CPT × CG should be transferred to the culture medium 35 days after pollination. Based upon this observation, crossed ovaries were collected 35 days after pollination, then ovules with placenta were explanted on culture medium and cultured in the dark at 25°C. Plantlets were induced from ovules cultured in MS medium containing 3% sucrose and 10% coconut milk. The hybrids (2n=66) derived from CPD × CG failed to yield viable seeds by self-pollination, although they showed some pollen fertility. In contrast, the hybrids (2n=90) derived from CPT × CG showed high pollen fertility and yielded viable seeds by self-pollination. Furthermore, they were resistant to disease caused by Fusarium oxysporum f. sp. cyclaminis, Erwinia herbicola pv. cyclamenae and Pseudomonas marginalis pv. marginalis.Abbreviations CPD C. persicum diploid - CPT C. persicum tetraploid - CG C. graecum