Investigations on F1 and F2 hybrids between Brassica oleraceae Var. Acephala and Raphanus sativus

A cross was made between B. oleraceae var. acephala and R. sativus. The use of a piece of cotton soaked with the growth regulator, N-m-tolyphthalamic acid at 100 ppm and attached to the pedicel at the time of pollination resulted in set of several pods with one viable seed. The hybrid kale x radish appeared vegetatively intermediate between the parents except for the flower color which resembled the radish. The F₁ plant propagated asexually and maintained under screen isolation cages with pollinating insects for a period of four years yielded only one viable F₂ seed. The single F₂ plant obtained, an allotetraploid, failed to yield any fertile seeds through selfing. Backcrossing of this F₂ plant to the radish has yielded two seeds.Department of Horticulture, Michigan State UniversityWith 7 figuresJournal article No. 2923. From the Michigan Agricultural Expreriment Station.     

Productivity of F hybrids of botanical varieties ofBrassica oleracea L.

Summary F₁ hybrids were obtained by making pair-crosses between nine botanical varieties ofBrassica oleracea. Although non-inbred parents were used, their yield was surpassed by all F₁ hybrids. The highest yields were shown by the crosses marrow-stem kale × sprouting broccoli and marrow-stem kale × cauliflower while the highest consumable yield (assessed by feeding plants to sheep) was that of F₁ hybrids marrow-stem kale × cauliflower and wild cabbage × cauliflower. A scheme for the production of F₁ hybrid seed is suggested, based upon an adaptation ofThompson's (1964) Triple-cross technique.     

RAPD markers linked to microspore embryogenic ability in Brassica crops

In order to identify the markers linked to microspore embryogenic ability in Brassica crops, RAPD segregation analyses were performed in a microspore-derived (MD) population and a F₂ population derived from F₁between ‘Ho Mei’ (high responsive parent in microspore embryogenesis) and ‘269’ (low responsive parent) in Chinese cabbage, and between ‘Lisandra’ (high responsive parent) and ‘Kamikita’ (low responsive parent) in oil seed rape. After 230 and 143 primers were screened, a total of 148 and 52markers were detected to be polymorphic between the parents in Chinese cabbage and oilseed rape, respectively. Twenty-seven percent of the markers in the MD population showed a significant segregation distortion in both crops. Of the markers showing segregation distortion in the MD population, 71–75% of the markers followed the expected Mendelian segregation ratio in the F₂ population. When the relationships between such distorted markers and microspore embryogenesis of the F₂ population were examined, 7 and 3 markers were identified to be associated with embryogenic ability in Chinese cabbage and oilseed rape, respectively. These markers showed additive effects on embryo yields, and the plants having more alleles of the high responsive parent produced higher embryo yields. These markers maybe useful in marker-assisted selection for improving microspore responsiveness straits in Brassica crops. This revised version was published online in July 2006 with corrections to the Cover Date.     

Efficiency of PCR-RF-SSCP marker production in Brassica oleracea using Brassica EST sequences

Efficiencies of SCAR, CAPS and PCR-RF-SSCP marker production were investigated using two combinations of breeding lines in Brassica oleracea. Published EST sequences of B. oleracea, Brassica rapa, Brassica napus, and Arabidopsis thaliana and newly determined nucleotide sequences of anther cDNA clones from B. oleracea were used for designing primer pairs to amplify genes. The percentage of primer pairs yielding DNA amplification of a single gene was higher in primer pairs of B. oleracea (91%) than those of B. rapa (56%) and A. thaliana (17%). Single DNA fragments amplified by 9% of the primer pairs showed polymorphism as SCAR markers between a broccoli line and a Chinese kale line by agarose-gel electrophoresis. CAPS analysis showed different band patterns in 32% of the same-sized DNA fragments, and PCR-RF-SSCP analysis revealed DNA polymorphism in 52% of those showing no DNA polymorphism by CAPS. In total, 71% of the single DNA fragments were converted to DNA markers. The frequency of DNA polymorphism between parental lines of a cabbage F₁ hybrid was lower, 5% by SCAR and 12% by CAPS. However PCR-RF-SSCP analysis revealed DNA polymorphism in 21% of the DNA fragments showing no polymorphism by CAPS. These results suggest that PCR-RF-SSCP analysis enables highly efficient DNA marker production for mapping of genes in Brassica using progeny, even progeny of closely related parents. Analysis of selfed seeds of broccoli F₁ cultivars using PCR-RF-SSCP markers indicated that PCR-RF-SSCP analysis is also applicable to seed purity tests.     

Transfer of resistance to race 2 of Plasmodiophora brassicae from Brassica napus to cabbage (B. oleracea ssp. capitata). III. First backcross and F2 progenies from interspecific hybrids between B. napus and B. oleracea ssp. capitata

Summary The first backcross and F₂ progenies from triploid F₁ and tetraploid F₁ hybrids between B. napus and 2x and 4x B. oleracea ssp. capitata (cabbage) were studied for their general morphology, resistance to race 2 of the clubroot pathogen, chromosome number and meiotic chromosome behavior. No linkage was apparent between resistance and the major morphological characters. Unreduced gametes played a large part in the successful formation of seed of the B₁ and F₂ progeny. B₁ plants with low chromosome numbers were selected for use in recurrent backcrosses. The potential use of anther culture to extract gametic progenies from resistant B₁ and F₂ plants with higher chromosome numbers was suggested. The presence of homoeologous pairing observed in all the plants is considered advantageous for selecting suitable progeny in later generations.     

Inheritance of seed colour in Brassica campestris L. and breeding for yellow-seeded B. napus L.

Summary Seed colour inheritance was studied in five yellow-seeded and one black-seeded B. campestris accessions. Diallel crosses between the yellow-seeded types indicated that the four var. yellow sarson accessions of Indian origin had the same genotype for seed colour but were different from the Swedish yellow-seeded breeding line. Black seed colour was dominant over yellow. The segregation patterns for seed colour in F₂ (Including reciprocals) and BC₁ (backcross of F₁ to the yellow-seeded parent) indicated that the black seed colour was conditioned by a single dominant gene. Seed colour was mainly controlled by the maternal genotype but influenced by the interplay between the maternal and endosperm and/or embryonic genotypes. For developing yellow-seeded B. napus genotypes, resynthesized B. napus lines containing genes for yellow seed (Chen et al., 1988) were crossed with B. napus of yellow/brown seeds, or with yellow-seeded B. carinata. Yellow-seeded F₂ plants were found in the crosses that involved the B. napus breeding line. However, this yellow-seeded character did not breed true up to F₄. Crosses between a yellow-seeded F₃ plant and a monogenomically controlled black-seeded B. napus line of resynthesized origin revealed that the black-seeded trait in the B. alboglabra genome was possibly governed by two independently dominant genes with duplicated effect. Crossability between the resynthesized B. napus lines as female and B. carinata as male was fairly high. The sterility of the F₁ plants prevented further breeding progress for developing yellow-seeded B. napus by this strategy.     

Variation of flowering time in inbred brussels sprouts and cabbage (Brassica oleracea L.)

Summary Differences in time of flowering between two parent lines of F₁ varieties of Brassica oleracea could significantly affect both the total seed yield and the proportion of sibs in the hybrid seed crop.The results for the time to flower of an incomplete diallel show that even in relatively inbred material of brussels sprouts there is potential to select for altered flowering time. Results obtained from a set of highly inbred cabbage lines which were given three temperature regimes in early spring show that between line variation is complicated by different responses of some of the lines to the environments.     

Interspecific hybridization between Brassica carinata and Brassica rapa

The crossability between Brassica carinata (BBCC, 2n=34) and Brassica rapa (AA, 2n=20), and the cytomorphology of their F₁ hybrids were studied. Hybrids between these two species were only obtained when B. carinata was used as the female parent. The hybrid plants exhibited intermediate leaf and flower morphology, and were found to be free from white rust and Alternaria blight diseases. One of the four F₁ plants was completely male sterile, while the remaining plants had 4.8, 8.6, and 10.9% stainable pollen, respectively. No seed was produced on hybrid plants under self pollination or in backcrosses; but seed was obtained from open pollination. The occurrence of the maximum of 11 bivalents as well as up to 44.8%) of cells with multivalent associations in the form of trivalents (0‐2) and a quadrivalent (0‐1) in the trigenomic triploid hybrid (ABC, 2n = 27) revealed intergenomic homoeology among the A, B and C genomes. Meiotic analysis of F₁ hybrids indicated that traits of economic importance, such as disease resistance, could be transferred from B. carinata to B. rapa through interspecific crosses.     

Morphological and molecular characterization of the second backcross progenies of Ogu-CMS Chinese kale and rapeseed

Ogura cytoplasmic male sterility (Ogu-CMS) is widely used in the production of commercial hybrids of Brassica oleracea. However, the widespread application of the Ogu-CMS system in B. oleracea has hindered the germplasm innovation of Ogu-CMS resources due to the lack of a natural restorer line. Previously, the Ogu-CMS fertility-restored interspecific hybrids between rapeseed 15Y403 (2n = 38, AACC) and Chinese kale JL1 (2n = 18, CC) have been successfully produced. However, these progenies, which still contained a large proportion of rapeseed genomic components, showed poor fertility and a low seed setting rate under natural pollination. To improve fertility and seed setting, a successive backcross with JL1 was performed to produce BC₂ progenies. Screening with the Rfo-specific marker, five individuals harboring the Rfo gene were identified among 98 BC₂ progenies. These five individuals underwent background marker screening and an evaluation of agronomic traits and fertility. One individual (code: 15Q23) was identified with higher pollen viability, better seed setting under natural pollination, and a closer genetic background to the parent Chinese kale JL1. Many morphological traits showed no significant differences (P < 0.05) between 15Q23 and the backcross parent JL1. However, the average seed setting of 15Q23 under natural pollination was 0.72 seeds per pod, which was 50 times higher than that of BC₁ progenies, and the average pollen viability was 87.07%, which was significantly better than that of the F₁ and BC₁ plants (P < 0.01). The genetic background of 15Q23 was more closer to the parent JL1 than that of BC₁ plants and another BC₂ fertility-restored individual, with 82% of the polymorphic alleles being the same as those of the parent Chinese kale JL1. Thus, the individual 15Q23 could be used as a donor plant for further backcrosses. This study lays the foundation for the development of Ogu-CMS restorer material in B. oleracea.     

An S-allele survey of cabbage (Brassica oleracea var. Capitata)

Summary A total of 31 S-alleles was found in a survey of 197 cabbage plants representing 11 cultivars of diverse type. Most of these S-alleles also occurred in either kale or Brussels sprouts, but five of them have not been found previously and apparently occur only in cabbage. A more detailed study of five cultivars of spring cabbage showed only 12 S-alleles in all, with 6–10 S-alleles in four older cultivars and only 3 S-alleles in the newer more highly selected cultivar. S2 was by far the commonest S-allele, as it is in B. oleracea as a whole. The highly recessive alleles S5 and S15 were not particularly common in cabbage and this may partly explain why the sib problem in F₁ hybrids is apparently less in cabbage than in Brussels sprouts. Three cases were found in which an S-allele was completely recessive in both the stigma and the pollen. The problems for the breeder created by this rather unusual situation are discussed.     

Inbreeding and the production of commercial F1 hybrid seed of brussels sprouts

Inbred lines, many of which have reached or passed the I₄ generation, have been produced by successive generations of self-pollination of material selected stringently for desirable characteristics. Of 99 parent plants originally selected from three different non-inbred varieties only eight were represented in the I₄ generation, by which time individual inbred lines were virtually true breeding for characters of commercial importance.An F₁ hybrid named Avoncross, produced by crossing unrelated selected inbred lines, has given consistently good yields of high quality sprouts in trials grown during several seasons. In most of these trials Avoncross gave the greatest marketable yield as compared with various control varieties and its superiority was significant at the 5% level of probability. A further F₁ hybrid, as yet un-named, has also given promising results in recent trials.Genetic male sterility was found in the material but did not provide an economic method for the production of F₁ hybrid seed. The method developed for the field production of F₁ hybrid seed relies on natural crossing between partially sib-compatible inbred lines. The partial sib-compatibility makes it possible to multiply the parent inbred lines by using natural sib-pollination in the field, but leads to the presence of sibs in the F₁ hybrid seed produced by natural crossing. The percentage of sibs in field crops of the F₁ hybrid established from such seed, however, was acceptably low because the majority of the sib-seedlings initially present could be rejected, on the basis of their smaller size, at the time of transplanting from the seedbed to the field.Experimental seed production, both of parent inbred lines and of the F₁ hybrid Avoncross, indicated that commercial production of F₁ hybrid seed in this manner was likely to be economically feasible.     

Assessment of DNA markers for seed contamination testing and selection of disease resistance in cabbage

Cabbage (Brassica oleracea L. var. capitata) is an important vegetable worldwide. Most Japanese commercial cultivars of cabbage use an F₁ hybrid seed production system. The purity of F₁ hybrid seeds is important and the assessment of purity based on DNA markers can be highly accurate. In addition, selection of agronomically important traits such as disease resistance based on DNA markers is useful for breeding of cabbage. The aim of this study is to demonstrate the effectiveness of DNA marker-assisted selection in cabbage. In this study we distinguished the parental S haplotypes in 35 F₁ hybrid cultivars by combining several linked DNA markers. Thirty-one highly polymorphic simple sequence repeats (SSR) markers were screened from 175 reported SSR markers, which are useful for assessment of the purity of F₁ hybrid seeds. We examined the relationship between the DNA marker based genotype and the phenotype by an inoculation test of clubroot disease. A co-dominant PCR–RFLP marker was developed for selection of Fusarium yellows resistance and the genotypes using this marker were consistent with inoculation test in all tested samples.     

Distribution of S haplotypes in commercial cultivars of Brassica rapa

Self‐incompatibility in Brassicaceae plants is sporophytically controlled by a single multi‐allelic locus (S locus), which contains at least three highly polymorphic genes expressed in the stigma (SLG and SRK) and in the pollen (SCR/SP11). Using polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) analysis with SXG‐specific primer pairs, the S haplotypes of F₁ hybrid and open‐pollinated commercial cultivars of Brassica rapa were identified. The number of S haplotypes detected in the F₁ hybrid cultivars of Chinese cabbage, komatsuna, pak‐choi, turnip, open‐pollinated cultivars of Chinese cabbage and turnip were 9, 9, 4, 11, 13 and 12, respectively. Nine of them had different PCR‐RFLP profiles from those of the S‐tester lines that determined the SLG sequences. Four SLG sequences in the F₁ hybrid cultivars were determined and named S⁵³, S⁵⁴, S⁵⁵ and S⁵⁶, respectively. It is demonstrated that the PCR‐RFLP analysis using specific primer pairs of SLG and SRK is useful for identification of the S haplotypes, in both, S homozygous and S heterozygous plants of B. rapa. The possibility of using this method routinely in breeding programmes, and in the evaluation of F₁ hybrid seed purity, is discussed.     

The transfer of triazine resistance from Brassica napus L. to B. oleracea L. III. First backcross to parental species

Summary Atrazine resistant Brassica napus × B. oleracea F₁ hybrids were backcrossed to both parental species. The backcrosses to B. napus produced seeds in both directions but results were much better when the F₁ hybrid was the pollen parent. Backcrosses to B. oleracea failed completely but BC₁s were rescued by embryo culture both from a tetraploid hybrid (2n = 4x = 37; A₁C₁CC) and sesquidiploid hybrids (2n = 3x = 8; A₁C₁C). Progeny of crosses between the tetraploid hybrid and B. oleracea had between 25 and 28 chromosomes. That of crosses between the sesquidiploid hybrid and B. oleracea had between 21 and 27. A few plants that had chromosome counts outside the expected range may have originated from either diploid parthenogenesis, unreduced gametes or spontaneous chromosome doubling during in vitro culture. Pollen stainability of the BC₁s ranged from 0% to 91.5%. All the BC₁s to B. oleracea were resistant to atrazine.     

Inheritance of cabbage head formation in crosses of cabbage × ornamental cabbage and cabbage × kale

We investigated the inheritance of head formation in Brassica oleracea by using two crosses, cabbage × ornamental cabbage and cabbage × kale. The degree of head formation (DHF) was classified into nine grades ranging from non‐heading to full heading. DHF in the two F₂ populations showed a continuous distribution. The variance of F₃ offspring selected for full heading or non‐heading was large. The DHF distributions in the F₃ offspring selected for moderate heading or randomly selected F₃ populations were similar to those of the respective F₂ populations, but had smaller average values and variance. However, the realized heritability in F₃ offspring was similar in full‐heading and non‐heading selections. Our findings suggest that head formation is a quantitative trait controlled additively with low dominance effects. In comparisons of leaf developmental patterns among the parents, only cabbage showed a change in leaf shape becoming wider because of the shorter petiole length with increasing leaf position. These findings suggest that cabbage acquired the developmental changes in leaves required to form a head during the process of domestication.     

Giemsa N-banding pattern in cabbage and Chinese kale

Summary In cabbage (Brassica oleracea var. capitata) and Chinese kale (B. oleracea alboglabra) four types of N-bands can be distinguished: pericentromeric, telomeric (terminal), intercalary and satellite bands. Typical NOR bands were not observed. The pericentromeric bands appear at the pericentric regions, possibly even at the centromeres of all chromosomes. Telomeric bands are observed on the short arms of chromosomes 1,5 and 6 in cabbage and chromosomes 1 and 5 in Chinese kale. Intercalary bands stained weakly in the long arms of chromosome 3 in cabbage and chromosome 2 in Chinese kale. Satellite bands cover the entire satellites in both Brassica species. The N-banding pattern is very similar in appearance to the C-banding pattern in both species and much more convenient to apply.     

Hybridization of Sinapis alba L. and Brassica napus L. via Embryo Rescue

Embryo rescue techniques were used to obtain hybrids between Sinapis alba L. (white mustard) and Brassica napus L. (oilseed rape) with the goal of improving the disease tolerance of oilseed rape. Hybrid plants with 31 or 43 chromosomes were only recovered, when S. alba, was used as the female parent. One hybrid was obtained from the cross S. alba L. cv. ‘Kirby’×B. napus L. cv. ‘Topas’, while 26 hybrids were obtained, when various S. alba L. cultivars were pollinated with the rapid cycling B. napus line CrGC 5006. All F₁, hybrid plants were male sterile; however, the first generation backcross to B. napus L., also obtained by embryo rescue, produced plants with 50 chromosomes and 61–84 % pollen viability. Second backcross generation seed was produced by normal sexual crossing. Preliminary cytological analyses of pollen mother cells of hybrid plants suggests the possibility of genetic exchange between the two species.     

Chasmogamous mutant,a novel character enabling commercial hybrid seed production in mungbean

Mungbean (Vigna radiata (L.) Wilczek) is a self-pollinating crop that displays significant hybrid vigor in seed yield of F₁ hybrids. Thus there is the possibility to use hybrid varieties as a breakthrough to raise the yield plateau of mungbean. However, hybrid mungbean seeds can only be accomplished by hand-pollination and thus commercial production is not possible. To encourage hybrid seed set, the plant breeder needs to develop characters that promote higher outcrossing rate such as open flower (chasmogamy). In this experiment, new chasmogamous mutants were induced by gamma irradiation at the rate of 100 and 200 Gy. The mutants were identified at a low rate of 0.4–0.7% in the M₂ generation of accession V1197, and observed for their purity by growing in plant-to-row in the M₃ and M₄ generations. A uniform chasmogamous line was hybridized to normal flower lines to study the inheritance of this character. All F₁ plants had normal flowers, while the F₂ plants segregated well with 3 normal : 1 chasmogamous ratio. When the F₁ was backcrossed to the chasmogamous parent, the progeny gave a ratio of 1 normal to 1 chasmogamous. Thus, chasmogamy was controlled by a single recessive gene, cha.     

Transfer of resistance to race 2 of Plasmodiophora brassicae from Brassica napus to cabbage (B. Oleracea spp. Capitata). IV. A resistant 18-chromosome B1 plant and its B2 progenies

Summary The somatic karyotype and meiotic chromosome behavior were studied in an 18-chromosome B₁ plant derived from backcrossing a triploid (Brassica napus x B. oleracea ssp. capitata) F₁ hybrid to cabbage. It is considered that cabbage chromosomes no. 1 and no. 7 were substituted by two shorter B. napus chromosomes. Meiotic disturbances were more apparent during the late stages of second division. Seed fertility of this plant was largely restored in the second backcrosses with both cabbage and broccoli. 18-chromosome B₂ plants resistant to race 2 of Plasmodiophora brassicae were recovered among the progenies.Contribution no. J. 725 from the Research Station, Research Branch, Agriculture Canada, St-Jean, Québec J3B 6Z8.     

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.