Introgression of Cajanus platycarpus genome into cultivated pigeonpea, C. cajan

Summary Cajanus platycarpus, an incompatible wild species from the tertiary gene pool of pigeonpea (C. cajan (L.) Millspaugh), has many desirable characteristics for the improvement of cultivated varieties. To necessitate such transfers, embryo rescue techniques were used to obtain F₁ hybrids. The F₁ hybrids were treated with colchicine to obtain tetraploid hybrids, that were selfed to obtain F₂, F₃ and F₄ progenies. All of the hybrids and subsequent progenies had an intermediate morphology between the two parents. Backcrossing of the tetraploid hybrids with cultivated pigeonpea was not possible given embryo abortion, with smaller aborted embryos than those obtained in the F₀ parental cross.As a route of introgression, diploid F₁ hybrids were backcrossed with cultivated pigeonpea and BC₁ progeny obtained by in vitro culture of aborting embryos. BC₂ plants were obtained by normal, mature seed germination. Although embryo rescue techniques had to be used to obtain F₁ and BC₁ plants, it was possible to produce BC₂ and subsequent generations through direct mature seed. Every backcross to cultivated pigeonpea increased pollen fertility and the formation of mature seeds.Special project assistant till December, 2003.     

Chromosome 1D as a possible location of a gene (s) controlling variation between wheat (Triticum aestivum L.) varieties for carbon isotope discrimination (Δ) under water-stress conditions

Cytoplasmic male-sterility (CMS) in pigeonpea has been reported when some wild relatives of pigeonpea were crossed as the female parent with cultivated types as the male parent. In this paper we report a new source of CMS developed by using the cultivated pigeonpea as the female parent and one of its wild relative Cajanus acutifolius as the pollen donor. This is the first report in pigeonpea where CMS has been developed using the cytoplasm of cultivated pigeonpea. Several pure line cultivars of pigeonpea restored pollen fertility whereas cv. HPL 24 partially maintained male-sterility. The wild species C. acutifolius used as one of the parents, maintained complete sterility. Cytological analysis revealed that both in male-sterile as well as the fertile floral buds, meiosis proceeded normally till the tetrad stage. However in the male-sterile genotypes during the formation of tetrads, the pollen mother cell (PMC) wall did not dissolve to release the tetrads unlike in the fertile genotypes and this major event was found to be responsible for male-sterility.     

Cross compatibility between Abelmoschus esculentus and A. moschatus

Interspecific cross compatibility between cultivated and wild okra (Abelmoschus esculentus and A. moschatus) and pollen tube growth behaviour in the crosses among a local cultivar of A. esculentus, A. moschatus and their F₁s were studied. Fruit set was observed in all the crosses except one and seed setting was absent in two of the crosses which set fruit. All seed produced were shrivelled but F₁ plants were obtained from two crosses where cultivated okra was used as the seed parent. The F₁ plants were perennial in nature with very low pollen viability and seed set. A high percentage of pollen germination and profuse pollen tube penetration in the style were observed in the cross A. esculentus × A. moschatus but low pollen tube penetration with abnormal pollen tubes was observed in the reciprocal cross. The number of pollen tubes was very low but they appeared to be normal in the backcross A. esculentus × F₁, but were generally abnormal in the reciprocal cross. Both pre- and postzygotic barriers seemed to occur in crosses between the two species. The present studies indicate that these barriers can be overcome and desirable characters from A. moschatus transferred to cultivated okra using conventional hybridisation techniques. This revised version was published online in July 2006 with corrections to the Cover Date.     

Production of hybrids betweenCajanus platycarpus andCajanus cajan

Summary Cajanus platycarpus a wild species of pigeonpea incompatible with the cultigen has many desirable characters important for the improvement of cultivated pigeonpea. In the present study, barriers to hybridization were studied and were identified as post-zygotic. Efficient embryo rescue techniques were developed. As a result, hybrids were produced. Morphological and cytological studies as well as esterase isozyme band pattern confirmed the hybrid nature of the plants. The F₁ hybrids were completely pollen sterile. Meiotic studies were carried out to check for the cause of pollen sterility.Abbreviations MS Murashige & Skoog's (1962) medium - BAP 6-Benzylaminopurine - IBA Indole-3-butyric Acid - NAA Naphthaleneacetic Acid     

Barriers to hybridization of Solanum bulbocastanum Dun. and S. Verrucosum Schlechtd. and structural hybridity in their F1 plants

Summary Results of reciprocal crosses between Solanum verrucosum (2n=2x=24) and S. bulbocastanum (2n=2x=24) are described in terms of pollen tube behaviour in styles, of berry and seed set, of fertility and of meiotic behaviour of the F₁ hybrids. Pollen tube growth of S. verrucosum is strongly inhibited in styles of S. bulbocastanum, whereas no inhibition is observed in the reciprocal cross. Therefore S. bulbocastanum x S. verrucosum fails to set berries or seeds, whereas the reciprocal cross produces both berries (54.4% berry set) and seeds (0.3 per berry). Only 14.6% of the seeds germinate. Both the diploid and corresponding tetraploid hybrid plants are vigorous, flower abundantly, have a rather regular meiosis (mainly rod bivalents), but show a high degree of cytoplasmic-genic male sterility. Crossability of the diploid hybrid plants is nil when used as pollen parents and near to zero when used as pistillate parents. The barriers to hybridization of the parent species (unilateral inhibition of pollen tube growth, somatoplastic sterility, cytoplasmic-genic male sterility and structural differences of the parental chromosomes) are discussed and methods are suggested to overcome these barriers.     

Variation in progenies of an Arachis hypogaea x diploid wild species hybrid

Summary Derivatives of a cross between cultivated peanuts, Arachis hypogaea L. (2n=40), and the wild species collection GKP 10017 (2n=20) were compared morphologically, for leafspot resistance and for yield. The objective of the study was to determine the effects of wild species germplasm on the A. hypogaea genome. The sterile F₁ hybrid which resulted from crossing the two species was treated with colchicine to restore fertility at the 6x ploidy level. The resulting hexaploid was cytologically unstable and progeny lost chromosomes until stability was regained at the 2n=40 chromosome level. Forty-seven characters were used to analyze the variation among plants in the tetraploid interspecific hybrid population. The plants were compared to four cultivated lines plus GKP 10017. Many hybrids were intermediate to the two parents in morphology. Individual traits such as growth habit, pod and seed size, elongation of the constricted area between pods, nodulation and leaflet size were altered by the presence of GKP 10017 germplasm in many of the hybrid plants. Cercospora arachidicola Hori and Cercosporidium personatum (Berk. & Curt.) Deighton resistances were evaluated for all plants. Several hybrids had few lesions due to either leafspot pathogen. In addition, 24 largeseeded interspecific hybrid selections were compared to the cultivated variety NC 5 for yield. Five selections were superior to both parents at p=0.01. Morphology, disease resistance and yields appeared to be greatly influenced by the wild species GKP 10017 germplasm in plants of the interspecific hybrid population. The potentials of using wild species for improvement of the cultivated peanut are discussed.Paper number 5948 of the journal series of the North Carolina Agricultural Research Service, Raleigh, NC 27650. The investigation was supported in part by ICRISAT and SEA-CR grant no. 701-15-51.     

Cytogenetic analysis of interspecific hybrids in genus Citrullus (Cucurbitaceae)

Investigations on interspecific crossability in two species of the genus Citrullus were studied based on their morphology, chromosome pairing and pollen fertility in their hybrids resulting from 12 combinations(including reciprocals). These have been utilized for tracing the phylogenetic relationship between the taxa and factors responsible for their differentiation. A collective evaluation of data on cross compatibility,morphology, chromosome association and pollen fertility indicated a close phylogenetic relationship between the species C. colocynthis (wild2n = 22) and C. vulgaris (cultivated 2n = 22). Significant differences for crossability and pollen fertility were observed between hybrids of C. colocynthis × C. vulgaris and reciprocals. Meiotic analysis of F₁ hybrids revealed that these two species are freely crossable and provide normal chromosome association, involving an average of 9 to maximum 11 pairs of chromosomes. This indicated the possibility of high recombination and thereby genetic introgression of desirable traits. As both the species are of economic significance, breeding strategies can be developed for the genetic improvement of these species. This revised version was published online in August 2006 with corrections to the Cover Date.     

Restoration of meiotic regularity and fertility among derivatives of Lens culinaris x L. Nigricans hybrids

Summary The wild lentil, L. nigricans is cross compatible with the cultivated species. Chromosome pairing in their F₁ hybrids is highly irregular due to a difference of three translocations between the parental species. However, the hybrids are partially fertile and in the F₂ populations there is a marked improvement in fertility. About 19% of the F₂ plants were meiotically stable, fully fertile and their chromosome arrangement was similar to that of the cultivated species. All these are clear indications for the possibility of rapid introgression between the two species and hence systematic screening of L. nigricans for traits of economic value is fully justifiable.     

Possibility of genetic improvement of pigeonpea (Cajanus cajan (L.)Millsp.) utilizing wild gene sources

Summary Various wild relatives of pigeonpea,Cajanus cajan, namely some species ofAtylosia andRhynchosia, possess desirable characteristics that could be utilized for effecting genetic improvement of this crop. In total 73 cross combinations among two cultivars ofC. cajan and one accession each of eightAtylosia species and one ofRhynchosia were attempted. Twelve hybrids were obtained. Seven of these were analysed for F₁ fertility and their utility for agronomic improvement of theC. cajan. Fertility behaviour of the different F₁ hybrids varied and indicated that potential of gene transfer between the two genera,Atylosia andCajanus, was as good as within the genusAtylosia. From F₂ and F₃ families ofC. cajan × A. scarabaeoides andC. cajan × A. albicans, plants were selected with greater physiological efficiency and agronomic superiority. The prospects of transferring pod borer resistance and higher seed protein content from someAtylosia species to pigeonpea are discussed.     

Cajanus platycarpus (Benth.) Maesen as the donor of new pigeonpea cytoplasmic male sterile (CMS) system

Cajanus platycarpus, a distantly related wild species, was successfully crossed with cultivated pigeonpea using embryo rescue and tissue culture techniques. Advance generation lines showed a range of desirable characters including cytoplasmic male sterility. A range of pigeonpea cultivars restored fertility and was maintained by a few lines including cultivar ICPL 85010. Clasmogamous flowers were observed in the male sterile lines. In such flowers anthers did not form di-adlephous bundle. Cytological analysis revealed that meiosis proceeded normally till the tetrad stage in those anthers with pollen grains. After which many of the pollen grains turned sterile. In the anthers with pollen grains, dehiscence was not observed, thus creating functional sterility. In many other anthers, pollen mother cells (PMCs) were not formed at all, giving rise to sepalous anthers. In conclusion two mechanisms of male sterility existed, one was premeiotic, where PMCs did not form and in the second, although PMCs gave rise to pollen grains, they were either partially or totally sterile accompanied by non-dehiscence of anther wall.     

A cytoplasmic-nuclear male-sterility system derived from a cross between <Emphasis Type="Italic">Cajanus cajanifolius</Emphasis> and <Emphasis Type="Italic">Cajanus cajan</Emphasis>

Cytoplasmic-nuclear male-sterility is an important biological tool, which has been used by plant breeders to increase yields in cross-pollinated cereals and vegetables by commercial exploitation of the phenomenon of hybrid vigor. In legumes, no such example exists due to the absence of an economic way of mass pollen transfer from male to female parent. Pigeonpea [Cajanus cajan (L.) Millsp.], however, is a different legume where a moderate level of insect-aided natural out-crossing (25–70%) exists and it can be used to produce commercial hybrid cultivars, if an efficient and stable cytoplasmic-nuclear male-sterility (CMS) system is available. This paper reports the development of a stable CMS system (ICP 2039A), derived from an inter-specific hybrid of Cajanus cajanifolius, a wild relative of pigeonpea, with a cultivar ICP 11501. Using this genetic material, designated as the A₄ cytoplasm, a number of fertility restorers and maintainers have been developed. The best short-duration experimental pigeonpea hybrid ICPH 2470 produced 3205 kg ha⁻¹ grain yield in 125 days, exhibiting 77.5% advantage over the control cultivar UPAS 120. At present, all the important biological systems necessary for a successful commercial hybrid breeding program are available in pigeonpea and the package of this technology has been adopted by private seed sector in India for the production and marketing of hybrid varieties.     

Breeding tomato for pollen tolerance to low temperatures by gametophytic selection

Haploid selection for traits related to pollen cold tolerance in tomato was performed in segregating populations derived from a Lycopersicon esculentum × L. pennellii hybrid. BC₁ populations were obtained by combining normal and low temperature treatments on two stages of pollen development: pollen formation, and germination and pollen tube growth. F₁ hybrids were cultivated under low and normal temperatures and their pollen was used to pollinate L. esculentum plants at low and normal temperatures. The four BC₁ populations obtained were tested for the quality and quantity of pollen produced at low temperatures. The population obtained by cold treatment at both stages had a significantly improved pollen germination ability at low temperatures. The two other coldselected BC₁ populations showed no differences compared with the unselected BC population. A second cycle of pollen selection, corresponding to BC₂, was applied in order to test its persistence in the subsequent generations and the possibility to further improve the character. This second cycle showed no improvement although some plants retained the high pollen germination ability at low temperatures that was observed in the first cycle. Hence, gametophytic selection of some characters related with tomato pollen performance may be feasible, at least for the first cycle of selection.     

Interspecific hybrids and possible phylogenetic relations in grain amaranths

Summary Phylogenetic relations among the three species in grain amaranth need investigation to provide information for breeding experiments germplasm conservation efforts, and decision on evolutionary patterns in the grain types. Hybrid development from crosses between species was studied to find out genetic relationship between them. Interspecific crosses were made among Amaranthus hypochondriacus, A. caudatus and A. cruentus in the glasshouse. The F₁ plants were relatively easy to obtain but had low pollen fertility (10.3–15.1%) and low seed set. A few of these hybrids did not produce seeds. Only a few F₁ seeds obtained in crosses between A. cruentus and A. caudatus. All the F₁ plants from these crosses died at the seedling stage. Crosses between A. cruentus and A. hypochondriacus produced few seeds. Most of the F₁ plants obtained from the seeds died at the seedling stage with only four plants growing to maturity but were sterile. Based on hybrid development, it was suggested that A. hypochondriacus and A. caudatus were genetically closer than the other two combinations of species studied. A. cruentus seemed to be genetically closer to A. hypochondriacus than it was to A. caudatus.Research was supported by Grand No. AMA-KE-4-83-22. (CRG GRANT) from the National Academy of Science, U.S.A.     

A new source of cytoplasmic male sterility in sunflower

Summary Interspecific substitutions of the nucleus of Helianthus annuus (2n=34) cv. Saturn into the cytoplasm of H. petiolaris (2n=34) by successive backcrossing, resulted in progenies with indehiscent anthers containing white, rather than normal yellow, pollen. Further backcrossing by cv. Saturn failed to restore pollen shed, suggesting that the male sterility was cytoplasmic. In vivo germination tests of pollen from 23 such plants from eight BC₅ lines, indicated complete pollen sterility for 14 plants, but normal seed set, suggesting that female fertility was not affected. Meiosis in all plants examined was normal.Crosses between seven sources of pollen-fertility restorer, one collection of wild H. annuus, and an existing source of cytoplasmic male sterility, resulted in a high frequency of plants with normal pollen shed in all F₁ progenies. However, no normal pollen shed was evident in F₁ progenies for similar crosses between BC₅ male-steriles and three of the seven restorer sources, nor for the single wild H. annuus evaluated. The foregoing suggests that the backcross substitution lines are a new source of cytoplasmic male sterility. The inheritance of restoration of pollen shed was complex and not fully elucidated. Some data suggested that two independent, complementary, dominant genes were required, but others indicated two to three independent, dominant genes.     

Cytological relationships among Eumelilotus species analysed using cytological observations of interspecific hybrids obtained by in vitro culture

Interspecific hybrid plants between Melilotus altissima (yellow flowers) and either M. alba (white flowers) or M. taurica (white flowers) and between M. suaveolens (yellow flowers) and M. wolgica (white flowers) which were inviable because of heavy chlorosis, were grown successfully by in vitro culture and reached the flowering stage. Pollen fertility of hybrid plants between M. altissima and M. alba was about 46% whereas hybrids between M. altissima and M. taurica and between M. suaveolens and M. wolgica showed about 25% and 18% of pollen fertility, respectively. All F₁s did not set any selfed seeds. Observing the pairing of chromosomes at metaphase I in the hybrid plants, it was revealed that the genomes of M. altissima and M. alba differ by a reciprocal translocation. This result could account for the partial sterility of pollen in the F₁ hybrids. On the other hand, hybrid plants between M. altissima and M. taurica and between M. suaveolens and M. wolgica showed normal pairing of chromosomes though pollen fertility of the hybrids was very low, suggesting that low pollen fertility was controlled by sterility gene(s). These results demonstrate that M. altissima and 7 other species of the subgenus Eumelilotus share the same chromosomal constitution. The only exception, M. alba, differs from the other 8 species by a segmental interchange. This revised version was published online in July 2006 with corrections to the Cover Date.     

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 four genes for stable hybrid sterility and an epistatic QTL from a cross between Oryza sativa and Oryza glaberrima

To further understand the nature of hybrid sterility between Oryza sativa and Oryza glaberrima, quantitative trait loci (QTL) controlling hybrid sterility between the two cultivated rice species were detected in BC₁F₁ and advanced backcross populations. A genetic map was constructed using the BC₁F₁ population derived from a cross between WAB450-16, an O. sativa cultivar, and CG14, an O. glaberrima cultivar. Seven main-effect QTLs for pollen and spikelet sterility were detected in the BC₁F₁. Forty-four sterility NILs (BC₆F₁) were developed via successive backcrosses using pollen sterility plants as female and WAB450-16 as the recurrent parent. Seven NILs, in which the target QTL regions were heterozygous while the other QTL regions as well as most of the reminder of the genome were homozygous for the WAB450-16 allele, were selected as the QTL identification materials. BC₇F₁ for the seven NILs showed a continuous variation in pollen and spikelet fertility. The four identified pollen sterility QTLs were located one each on chromosomes 1, 3, 7 and 7. Pollen sterility loci qSS-3 and qSS-7a were on chromosomes 3 and 7, respectively, which coincides with the previously identified S19, and S20, while loci qSS-1 and qSS-7b on chromosomes 1 and 7L appear distinct from all previously reported loci. An epistatic interaction controlling the hybrid sterility was detected between qSS-1 and qSS-7a.     

Interspecific crosses in the genus Lycopersicum

The use of a tetraploid tomato as a pistillate parent in the crosses with Lycopersicum peruvianum and Lycopersicum glandulosum, instead of a diploid tomato, increases about 80 times the set of hybrid seed and thus gives a broader basis for the breeding work. The results seem to indicate that in the crosses on a diploid level, only unreduced eggcells of the tomato fertilized by haploid pollen of the above wild species, develop into normal seeds, and give rise to sesquidiploid F₁ progeny.     

Intergeneric hybrids and an amphiploid between Elymus canadensis and Psathyrostachys juncea

Summary Fifty four hybrid plants between Elymus canadensis and Psathyrostachys juncea were obtained by handpollination and embryo culture. The average cross compatibility between both species was 31.2 percent. One amphiploid plant was induced by colchicine treatment. The hybrid and amphiploid plants resembled P. juncea in appearance but showed a higher plant height and dry matter yield than the parents. The hybrids showed extremely low pollen stainability and were completely sterile. With the exception of one plant (2n=3x+1=22), all hybrid plants were allotriploids (SHN, 2n=3x=21). The amphiploid plant (SSHHNN, 2n=6x=42) showed 58.9% pollen stainability and 11.6% seed fertility.Mean chromosome associations of the hybrids and amphiploid at metaphase I were 0.02IV+0.06III+2.03II+16.91I and 0.07III+18.00II+5.85I, respectively. Lagging chromosomes, chromosome bridges, abnormal cytokinesis, and micronuclei were occasionally observed at the anaphase, telophase, or tetrad stage.     

Interspecific hybridization between Coffea arabica L. and tetraploid C. canephora P. ex Fr. I. Fertility in F1 hybrids and backcrosses to C. Arabica

Summary The interspecific (C. arabica × tetraploid C. canephora) F₁ hybrid showed a low but highly variable fertility. An almost complete restoration to normal fertility was observed in individual trees of BC₁ and BC₂ generations with C. arabica as recurrent parent. From a comparison of various tests of fertility, it was found that pollen viability is a satisfactory measure of fertility.