Interspecific hybridization in Cucumis (L.). I. Need for genetic variation,biosystematic relations and possibilities to overcome crossability barriers

Summary The genetic variation in pickling and slicing cucumbers (C. sativus L.) seems insufficient to enable the breeder to solve serious cultivation problems. Wild allies of C. sativus possess a wide range of interesting characters which could be incorporated by means of species crosses. In the past, attempts to achieve such crosses have failed. Important characters of Cucumis species are mentioned and biosystematic problems discussed. Results of previous research on species crosses prompted an examination of the prospects of further research.     

Methods of overcoming interspecific barriers in Trifolium

Summary Barriers to interspecific hybridization in Trifolium were investigated by manipulation of mentor pollen treatments, ploidy levels, and compatibility and male sterility systems. Crosses involving the addition of mentor pollen produced fewer seeds and hybrids than crosses involving normal pollination. Lower seed set with mentor pollen was deduced to result from the use of less viable pollen, approximately half the pollen having been killed by alcohol. Pollinations at the diploid level resulted in more hybrids than at the tetraploid level, perhaps because genes for male sterility produced higher female sterility in the tetraploids. The self-compatible stock produced more seeds, mostly selfs, than the self-incompatible stock, but produced more hybrids only in one cross, T. pratense L. × T. diffusum Ehrh. The use of male-sterile female parents reduced selfing but produced fewer hybrids than male-fertile female parents. Techniques of this study were designed to affect prefertilization barriers, but the lack of effect may indicate that postfertilization barriers in Trifolium are of greater importance.Journal Article No. 98-3-208 of Kentucky Agricultural Experiment Station. Published with approval of the Director.     

Synthesis and preliminary characterization of a new species (amphidiploid) in Cucumis

A successful interspecific hybridization between cucumber (Cucumis sativus L., 2n = 14) and a wild it Cucumis species, C. hystrix Chakr. (2n = 24) was made via embryo rescue. Hybrid plants (2n = 19; 7 from cucumber and 12 from C. hystrix) were sterile, but morphologically uniform. Self-pollination and backcrossing of F₁ hybrid plants to either parent confirmed presence of both male- and female-sterility that were likely caused by lack of homology and improper pairing during meiosis. While the multiple-branching habit, densely brown hairs (on corolla and pistil), orange-yellow corolla, and ovate fruit of F₁ hybrid plants were similar to that of the C. hystrix parent, the appearance of the first pistillate flower was more similar to that of C. sativus parent. The diameter and internode length of the stem, and the shape and size of leaves and flowers were intermediate when compared to the parents. The chromosome number in the hybrid was doubled through somaclonal variation during embryo culture and regeneration process to restore the fertility. Pollen grains were released and fruits with viable seeds matured on fertile, synthetic amphidiploid plants. The results from flow cytometry indicated that, on average, 7.3% of the morphologically unique regenerants had the 4C DNA content of 2.35 pg relative to the 2C DNA content of the original F₁ hybrid at 1.17 pg and, therefore, were likely chromosome-doubled F₁ hybrids (2n = 38). Nutrition alanalysis indicated that the synthetic species had higher protein (0.78%)and mineral (0.35%) content compared to the normal pickling cucumber(0.62% and 0.27%, respectively), and could be considered a new Cucumis crop having a special place in the future agriculture. Preliminarily evaluation indicates that C. hystrix possesses a high level of root-knot nematode resistance, and that this resistance is partially expressed in the interspecific F₁ and chromosome-doubled F₁. This and the fact that the fruit morphology of the fertile amphidiploid differs during the growing season (e.g., short and long fruit) suggest that it could be useful in broadening the germplasm base of cucumber. This revised version was published online in July 2006 with corrections to the Cover Date.     

Crossability relationships among Cajanus,atylosia and Rhynchosia species and detection of crossing barriers

Summary Crossability of two cultivars of Cajanus cajan, eight species of Atylosia and one of Rhynchosia was investigated. Of the 73 combinations attempted, success was achieved in 12 cases. C. cajan crossed successfully with A. albicans, A. cajanifolia, A. lineata, A. scarabaeoides, and A. trinervia. Within the genus Atylosia, A. lineata crossed with A. albicans and A. scarabaeoides, and A. scarabaeoides with A. sericea. Three species A. platycarpa, A. volubilis and R. rothii did not cross with any other one. In most of the unsuccessful combinations, although the pollen germinated on the receiving stigmas, the pollen tube growth was inhibited inside the stigma or in the stylar tissue.     

Interspecific hybridization in Cucumis L. II. The crossability of species,an investigation of in vivo pollen tube growth and seed set

Summary The crossability of 12 Cucumis species of African and Asiatic origin was studied in a diallel cross, in order to find ways to realise the cross between the common cucumber (C. sativus L.) and its wild relatives which carry resistances against diseases and pests.Self-pollinations and cross-pollinations within species gave normal pollen tube growth and seet set. The different accessions within a species, as a rule reacted alike in interspecific crosses. In crosses between African species different crossing patterns were found, viz. bilateral congruity, bilateral incongruity and unilateral incongruity. Within C. sativus all accessions intercrossed freely, except one, which displayed unilateral incongruity.Good seed was harvested from several crosses and in some cases embryo culture was needed for further development of seeds. No good seeds were obtained from any cross between a species of the African group and C. sativus L.     

Relationships between African species of the genus Cucumis L. Estimated by the production,vigour and fertility of F1 hybrids

Summary The production, vigour, and fertility of F₁ hybrids between nine African species of the genus Cucumis L. were studied as a measure of the relationships between the species. Hybrid plants were obtained from 29 out of the 72 possible cross combinations. Two F₁ hybrids died as seedlings, and 27 hybrids were raised to maturity. Pollen production and stainability varied greatly amongst these hybrids, as did fruit and seed set following self pollination and backcrossing with either parental species. The fruit shape of the hybrids was always intermediate between that of both parental species.Two species appeared to be closely related: C. prophetarum L. and C. anguria var. longipes A. Meeuse. Most other species produced highly to moderately fertile F₁ hybrids with at least one other species. C. metuliferus Naud. produced only sterile hybrids with C. zeyheri Sond. 2x. The results sustain the recent taxonomic classification of the genus (Jeffrey, 1980).     

Interspecific hybridization between rice bean (Vigna umbellata) and its wild relative (V. minima): Fertility-sterility relationships

Summary The pre-and post-fertilization barriers in the interspecific crosses between Vigna umbellata and V. minima were investigated. In the reciprocal crosses (V. minima as the parent) the entry of pollen tubes into the ovary was delayed by about 4 h, and no seed set was observed. However, no pre-fertilization barriers were encountered in crosses involving V. minima as the parent and V. umbellata as the parent (normal cross). The delay/absence of divisions in the endosperm and the failure of embryo to divide were the post-fertilization barriers responsible for somatoplastic sterility in normal crosses which yielded a few hybrid seeds. The hybrid seeds showed poor germinability. The F₁ hybrids were intermediate between the parents in most morphological characters, and are completely sterile for pollen. Backcrossing of F₁ hybrid with both the parents did not restore fertility in the progenies. V. minima is considered as the tertiary gene pool of the rice bean.     

Identification of barriers to interspecific crosses in the genus Trifolium

Summary A study of pre- and post-fertilisation barriers after interspecific crosses of diploid and tetraploid Trifolium pratense L. and wild species T. alpestre L., T. medium L. and T. sarosiense Hazsl. was aimed at finding of a promising cross combination for obtaining hybrids. The growth of pollen tubes was arrested in interspecific crosses mainly when T. pratense was at a diploid level. To investigate the post-fertilisation barriers in detail, the hybrid embryo viability was traced by two clearing treatments of immature seeds: (1) using chloral hydrate (which proved to be most appropriate); and (2) a mixture of benzyl benzoate and dibutly phthalate. In interspecific combinations T. pratense (4×) × either T. alpestre or T. sarosiense, enlargement of immature seeds occurred, but no hybrid embryo was traced. Of the wild species used as a male parent for crosses, T. medium was the only exception from the point of view of fertilisation. Globular, heart and the early torpedo stages of hybrid embryos were observed 7 days after pollination (DAP) but only when T. pratense was at a tetraploid level. When T. pratense (2×, 4×) was used as a male parent for interspecific crosses with T. alpestre, T. medium and T. sarosiense, strong defects in various stages of embryogenesis were observed, particularly wrinkled and narrowing embryo sacs caused by an expansion of endothelial cells. We conclude with the following finding: (1) to make crosses only in one direction with T. pratense as a female parent and T. medium as a male; (2) to use tetraploid plants of T. pratense; (3) and to excise hybrid embryos at an early torpedo stage, about 7 DAP.     

Assignment of Endosperm Balance Numbers to the tuber-bearing Solanums and their close non-tuber-bearing relatives

The Endosperm Balance Number (EBN) of over 80 species and subspecies of the tuber-bearing Solanums and their close non-tuber-bearing relatives representing 13 taxonomic series has been determined, with no species assigned to more than one EBN level. Among North American species, most diploids are 1EBN, most tetraploids are 2EBN and all hexaploids are 4EBN; however, among South American species most diploids are 2EBN, most tetraploids are 4EBN and again all hexaploids are 4EBN. Thus species may be isolated from others of the same ploidy level by EBN differences, e.g., 4×(2EBN) from 4×(4EBN), while other species differing in ploidy but having the same EBN may be intercrossed, e.g., 4×(2EBN) and 2×(2EBN). Chromosome doubling or 2n gametes can be used to make a lower EBN species compatible with a higher EBN species. These findings also explain the major crossing difficulties previously inherent in the use of North American species in potato improvement. They also have direct implications for potato improvement, barring the occurrence of other incompatibility barriers. Any 4×(4EBN) cultivar is endosperm compatible and thus will cross with 4×(4EBN) and 6×(4EBN) species. The 2×(2EBN) haploids of 4×(4EBN) cultivars likewise will hybridize with 2×(2EBN) and 4×(4EBN) species. All 2×(1EBN) species are crossable with 2×(2EBN) haploids through 2n gametes or chromosome doubling.Cooperative investigation of the U.S. Department of Agriculture, Agriculture Research Service, and the Wisconsin Experiment Station.     

Variation of prezygotic barriers in the interspecific hybridization between oilseed rape and wild radish

Escape of transgenes from genetically modified oilseed rape, Brassica napus, into wild radish, Raphanus raphanistrum, depends on sexual compatibility. The variation in prezygotic barriers of two different cultivars for interspecific hybridization with a population of wild radish was investigated by hand crossing and fluorescence microscopy of pistils. Significant differences were observed between oil seed rape cultivars in their ability to accept wild radish pollen germinating onto their stigma and the rate of fertilization of ovules. Some differences among the pollen donor plants were also detected. These results suggest that the rate of interspecific hybridization in the field would depend upon the oilseed rape cultivar and the genotype composition of the local wild populations. The implication of S-related genes, as revealed through identification by pistil tissue prints of class I and II S-types of SLG (S-Locus Glycoprotein) and SLR1 (S-Locus Related),and immuno-IEF, was not significant. This revised version was published online in August 2006 with corrections to the Cover Date.     

Polygon of crossability between eleven species of Ipomoea: section Batatas (Convolvulaceae)

Summary Crossability within Ipomoea section Batatas is complex because of genetic, cytogenetic and physiological interactions. This complexity is responsible for the fact that knowledge about phylogenetic relationships within this section remains preliminary. Between 1988 and 1991, studies of planting, pollination and evaluation of progenies were carried out at the facilities of CIP in La Molina and San Ramon, Peru. Self-compatibility was found in nine diploid species. Self-incompatibility was found in diploid I. trifida, tetraploid I. tiliacea and both tetraploid and hexaploid I. batatas. Sexual compatibility is though to be related to a multiallelic sporophytic incompatibility system which is expressed in the stigmatic papillae. Altogether, 4,162 cross pollinations were made between 11 species and 76 interspecific combinations of 110 possible combinations in a diallel 11×11 design. From these 76 interspecific combinations only 38 survived, and in these crossability (= proportion between number of fruits harvested/pollinations made) ranged from 0.01 to 1.00 at La Molina. At San Ramon, 11 out of 17 cross combinations were successful, and in these crossability ranged from 0.01 to 0.71. Crossability appeared to be influenced by latitude which is related to flowering synchrony. The early death of seedlings is hypothesized to be related to a genomic imbalance between embryo and endosperm. Maternal effects were found in crosses between diploids and tetraploids which produced triploid and diploid offspring, and in crosses between hexaploids and diploids which gave rise to hexaploid offspring. This study shows that I. trifida and I. x leucantha may act as bridge species for gene flow from wild Ipomoea species to the genepool of the sweetpotato.     

High regeneration potential in vitro of sunflower (Helianthus annuus L.) lines derived from interspecific hybridization

Successful selection of interspecific hybrid progenies with superior ability to regenerate shoots from apical meristems was performed in sunflower which now allows for the development of lines for improved biotechnological applications. Early generations of interspecific hybrids originating from crosses between the two H. annuus CMS lines ‘HA89’ and ‘Baso’, and 9 wild species were screened for their ability to regenerate in vitro. Evaluation of 36 progenies allowed to identify seven progenies from crosses involving H. mollis, H. giganteus, H. strumosus, and H. decapetalus which showed a significantly higher regeneration potential than the commercial hybrid ‘Albena’ regarding the number of shoots per explant. Among these progenies, 47.2 to 62.4% of explants produced shoots with an average of 2.3 to 3.5 shoots per cultured explant. Regeneration in vitro was significantly determined by the genotype. More than half of the investigated interspecific hybrids performed better than the inbred ‘HA89’ demonstrating that the high regeneration potential available in the wild species can be efficiently transferred to cultivated sunflower. The seven progenies with high regeneration potential in vitro were characterised by agronomic performance in the field. Two of the interspecific hybrids derived from H. strumosus and H. decapetalus not only showed a superior regeneration potential but also proved to be competitive to commercial hybrids with regard to important agronomic traits, e.g. fat content and TGW. This revised version was published online in July 2006 with corrections to the Cover Date.     

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.     

Overcoming unilateral breeding barriers between Lycopersicon peruvianum and cultivated tomato,Lycopersicon esculentum

Summary Treatment of Lycopersicon peruvianum stigmas with an artificial medium analogous to stigmatic exudate allowed pollen germination and growth on immature pistils. Growth of Lycopersicon esculentum pollen tubes to L. peruvianum ovules, an otherwise incompatible cross, was achieved following such treated bud-pollinations. No plantlets were recovered, although a few embryos from this cross at the globularity heart stage of development were excised at 22 days after pollination, indicating the presence of crossing failures as severe as in the reciprocal cross. Hybrid plants were obtained from the reciprocal cross, using as pollen parent an L. peruvianum line selected for congruity with L. esculentum. Bud pollinations to L. peruvianum, using these interspecific F₁ hybrids as the pollen parent, allowed viable embryo development and plantlet recovery. Resulting backcross plants, which possess approximately 1/4 L. esculentum genome in a L. peruvianum cytoplasm, may facilitate further introgression of the L. esculentum nuclear genes into in foreign cytoplasm.     

Interspecific hybridization between urdbean (Vigna mungo (L.) Hepper) and adzukibean (Vigna angularis (Willd.) Ohwi and Ohashi)

Summary To introduce resistance to fungal and viral diseases into urdbean (Vigna mungo), it was crossed with adzukibean (Vigna angularis) to achieve an interspecific hybrid between the two species by using different non-conventional breeding methods. Partial success was obtained with the use of UV irradiated pollen, intraspecific hybrids as parents and spray of immuno-suppressants on the female parent under glass house conditions and to the extent of callusing in aseptic culture.     

The production of multispecific octoploids from Fragaria species and the cultivated strawberry

Summary A method of incorporating genetic material from five species of Fragaria and the cultivated strawberry into fertile octoploids is described. A synthetic octoploid derived from a hexaploid x diploid hybrid was crossed to octoploid cultivar breeding lines until a BC₂ hybrid was produced. A second synthetic octoploid in which two diploid species and a tetraploid species were combined was crossed to a cultivated strawberry to produce a hybrid breeding clone. The two breeding clones were crossed and 222 seedlings were produced. The seedlings were generally vigorous and fruitful, some having commercial potential.     

Effects of gamma irradiation on in vitro pollen germination of different Cucumis species

Summary Pollen grains of 14 Cucumis accessions were irradiated with 0, 1, 2, 3 or 4 kGy acute gamma rays and germinated in vitro directly afterwards. Pollen germination was significantly reduced by increasing irradiation dose for all species, except C. melo var. agrestis. Pollen tube growth was generally reduced likewise. Pollen of two C. anguria subspecies was most sensitive to irradiation. Sensitivity of the pollen with respect to pollen tube growth in relation to irradiation dose was inversely related to total DNA amount per nucleus. In vitro germination was not related to DNA amount per nucleus. Results show that the examined Cucumis species, especially C. melo var. agrestis, are sufficiently resistant to irradiation to be used as donor species for in vivo egg cell transformation of the cucumber.     

The influence of genotype and environment on seed and embryo development in barley (Hordeum vulgare L.) after crossing with Hordeum bulbosum L.

Summary Seed and embryo development was studied in crosses between H. bulbosum and the barley genotypes VK 16032, Vada and Vogelsanger Gold and subsequently the F₁ and F₂ progenies from VK 16032 x Vada and VK 16032 x Vogelsanger Gold. Both seed and embryo development are strongly influenced by the barley genotype. Favourable environmental conditions can promote seed and embryo development in genotypes with existing good characters. Dominant inheritance for good seed development and incomplete dominance for large embryos is evident. Linkage between small embryo size and winter habit exists in Vogelsanger Gold.     

Genetic and genotype × environment interaction effects analysis for erucic acid content in rapeseed (Brassica napus L.)

Genetic main effects and genotype × environment (GE) interaction effects for erucic acid content (EAC) in rapeseed(Brassica napus L.) were analyzed for two year data by using the genetic models and corresponding statistical approaches for quantitative traits of diploid plant seeds. Eight parents were included in adiallel mating design in two environments. It was found that the embryo, cytoplasmic and maternal main effects and their GE interaction effects could simultaneously affect the performance of EAC, especially for the cytoplasmic and maternal effects. Since the amount of genetic main effects from the expression of genes in different genetic systems accounted for about 64.1%of the total genetic effects, EAC of rapeseed was mainly controlled by genetic main effects. The total narrow-sense heritability for EAC was 83.6% with the general heritability being 51.9% and the interaction heritability being 31.7%. It was suggested by the predicted genetic effects, that Tower and Youcai 601 were better than other parents for reducing EAC in rapeseed quality breeding. This revised version was published online in July 2006 with corrections to the Cover Date.     

Identification of reciprocal translocations observed in several Melilotus species (subgenus Eumelilotus) by interspecific triple crossings

Summary Melilotus alba differs by a reciprocal translocation from 7 other species which are categorized into M. officinalis and M. dentata groups. The two species groups, however, remained to be studied in relation to their cytological relations because of early degeneration of hybrid embryo. Interspecific triple crosses were successfully made in order to examine whether or not the reciprocal translocations observed were of the same origin. When F₁ hybrids between M. alba and M. officinalis group were crossed with M. dentata group, about a half of hybrids plants were heterozygous for a reciprocal translocation while the remaining plants were normal chromosome pairing, showing a segregation ratio of 1:1. This result indicates that reciprocal translocations observed among the three groups are of the same origin. Accordingly, it is expected that M. officinalis group and M. dentata group have the same chromosomal constitutions.