Mapping of seed quality traits in the C genome of Brassica napus by using a population carrying genome content of B. oleracea and their effect on other traits.

Increasing seed oil and protein contents and reducing the content of seed glucosinolates (GSLs) in Brassica oilseed crops are important objectives in breeding. By using an oilseed rape (B. napus L.) doubled-haploid (DH) population carrying genome content introgressed from Chinese kale (B. oleracea L.), we mapped quantitative trait loci (QTL) for these seed quality traits and investigated their effect on other traits including seed yield. A stable QTL for seed oil content was identified on chromosome C5 at 40-42 Mb position and a QTL for seed GSL content was identified on C9 at 7-8 Mb position. The C5 and C9 QTL alleles for high oil and GSL contents were derived from Chinese kale, demonstrating that high-oil QTL allele can be found in the parental species of oilseed rape. The low-GSL QTL allele of C9 exerted a significant positive effect on seed protein content, demonstrating that selection for this QTL allele contributed to higher protein content in canola seed. These two QTL were not affected by field environmental conditions and did not exert a significant effect on days to flowering and seed yield. Thus, the genomic regions and the molecular markers identified in this study should be useful in molecular breeding of the seed quality traits in oilseed rape.

Evaluation of Brassica oleracea accessions for resistance to Plasmodiophora brassicae and identification of genomic regions associated with resistance.

Clubroot disease caused by Plasmodiophora brassicae is a challenge to Brassica crop production. Breakdown of resistance controlled by major genes of the Brassica A genome has been reported. Therefore, identification of resistance in the Brassica C genome is needed to broaden the genetic base of resistance in Brassica napus canola. In this study, we evaluated 135 Brassica oleracea accessions, belonging to eight variants of this species to identify resistant accessions as well as to identify the genomic regions associated with resistance to two recently evolved P. brassicae pathotypes, F3-14 (3A) and F-359-13 (5X L-G2). Resistance to these pathotypes was observed more frequently in var. acephala (kale) followed by var. capitata (cabbage); few accessions also carried resistance to both pathotypes. Association mapping using single nucleotide polymorphism (SNP) markers developed through genotyping by sequencing technique identified 10 quantitative trait loci (QTL) from six C-genome chromosomes to be associated with resistance to these pathotypes; among these, two QTL associated with resistance to 3A and one QTL associated with resistance to 5X L-G2 carried ≥3 SNP markers. The 10 QTL identified in this study individually accounted for 8%-18% of the total phenotypic variance. Thus, the results from this study can be used in molecular breeding of Brassica crops for resistance to this disease.

Fine mapping of the major QTL for seed coat color in Brassica rapa var. Yellow Sarson by use of NIL populations and transcriptome sequencing for identification of the candidate genes.

Yellow seed is a desirable trait in Brassica oilseed crops. The B. rapa var. Yellow Sarson carry unique yellow seed color genes which are not only important for the development of yellow-seeded oilseed B. rapa cultivars but this variant can also be used to develop yellow-seeded B. napus. In this study, we developed near-isogenic lines (NILs) of Yellow Sarson for the major seed coat color QTL SCA9-2 of the chromosome A9 and used the NILs to fine map this QTL region and to identify the candidate genes through linkage mapping and transcriptome sequencing of the developing seeds. From the 18.4 to 22.79 Mb region of SCA9-2, six SSR markers showing 0.63 to 5.65% recombination were developed through linkage analysis and physical mapping. A total of 55 differentially expressed genes (DEGs) were identified in the SCA9-2 region through transcriptome analysis; these included three transcription factors, Bra028039 (NAC), Bra023223 (C2H2 type zinc finger), Bra032362 (TIFY), and several other genes which encode unknown or nucleic acid binding protein; these genes might be the candidates and involved in the regulation of seed coat color in the materials used in this study. Several biosynthetic pathways, including the flavonoid, phenylpropanoid and suberin biosynthetic pathways were significantly enriched through GO and KEGG enrichment analysis of the DEGs. This is the first comprehensive study to understand the yellow seed trait of Yellow Sarson through employing linkage mapping and global transcriptome analysis approaches.

Potential of the C Genome of the Different Variants of Brassica oleracea for Heterosis in Spring B. napus Canola.

The genetic base of Brassica napus canola need to be broadened for exploitation of heterosis at a greater level in the breeding of F1 hybrid canola cultivars. In this study, we evaluated 228 inbred B. napus canola lines derived from six B. napus × B. oleracea interspecific crosses and following two breeding methods (F2- and BC1-derived lines) to understand the effect of the B. oleracea alleles on heterosis for different agronomic and seed quality traits. Test hybrids of the inbreds derived from crosses involving vars. botrytis (cauliflower), alboglabra (Chinese kale) and capitata (cabbage) cv. Badger Shipper, on an average, gave about 10% mid-parent heterosis (MPH), and about 67% of the test hybrids gave higher seed yield than the common B. napus parent indicating that B. oleracea alleles can contribute to heterosis for seed yield in spring B. napus canola hybrids. This was also evident from a positive correlation of the genetic distance of the inbred lines from the common B. napus parent with MPH for seed yield (r = 0.31) as well as with hybrid yield (r = 0.26). Almost no correlation was found between genetic distance and MPH for seed oil and protein content as well as with the performance of the test hybrids for these two traits. The occurrence of positive correlation between seed yield of the inbred lines and test hybrids suggested the importance of the genes exerting additive effect for high seed yield in the hybrids. Very little or almost no heterosis was found for the other agronomic traits as well as for seed oil and protein content. While comparing the two breeding methods, no significant difference was found for seed yield of the test hybrids or the level of MPH; however, the BC1-derived inbred and test hybrid populations flowered and matured earlier and had longer grain-filling period than the F2-derived population. Thus, the results suggested that the B. oleracea gene pool can be used in the breeding of spring B. napus canola to improve seed yield in hybrid cultivars.

Molecular mapping of QTL alleles of Brassica oleracea affecting days to flowering and photosensitivity in spring Brassica napus.

Earliness of flowering and maturity are important traits in spring Brassica napus canola-whether grown under long- or short-day condition. By use of a spring B. napus mapping population carrying the genome content of B. oleracea and testing this population under 10 to 18 h photoperiod and 18 to 20 0C (day) temperature conditions, we identified a major QTL on the chromosome C1 affecting flowering time without being influenced by photoperiod and temperature, and a major QTL on C9 affecting flowering time under a short photoperiod (10 h); in both cases, the QTL alleles reducing the number of days to flowering in B. napus were introgressed from the late flowering species B. oleracea. Additive effect of the C1 QTL allele at 14 to18 h photoperiod was 1.1 to 2.9 days; however, the same QTL allele exerted an additive effect of 6.2 days at 10 h photoperiod. Additive effect of the C9 QTL at 10 h photoperiod was 2.8 days. These two QTL also showed significant interaction in the control of flowering only under a short-day (10 h photoperiod) condition with an effect of 2.3 days. A few additional QTL were also detected on the chromosomes C2 and C8; however, none of these QTL could be detected under all photoperiod and temperature conditions. BLASTn search identified several putative flowering time genes on the chromosomes C1 and C9 and located the physical position of the QTL markers in the Brassica genome; however, only a few of these genes were found within the QTL region. Thus, the molecular markers and the genomic regions identified in this research could potentially be used in breeding for the development of early flowering photoinsensitive B. napus canola cultivars, as well as for identification of candidate genes involved in flowering time variation and photosensitivity.

Construction of genetic linkage map and mapping of QTL for seed color in Brassica rapa.

A genetic linkage map of Brassica rapa L. was constructed using recombinant inbred lines (RILs) derived from a cross between yellow-seeded cultivar Sampad and a yellowish brown seeded inbred line 3-0026.027. The RILs were evaluated for seed color under three conditions: field plot, greenhouse, and controlled growth chambers. Variation for seed color in the RILs ranged from yellow, like yellow sarson, to dark brown/black even though neither parent had shown brown/black colored seeds. One major QTL (SCA9-2) and one minor QTL (SCA9-1) on linkage group (LG) A9 and two minor QTL (SCA3-1, SCA5-1) on LG A3 and LG A5, respectively, were detected. These collectively explained about 67% of the total phenotypic variance. SCA9-2 mapped in the middle of LG A9, explained about 55% phenotypic variance, and consistently expressed in all environments. The second QTL on LG A9 was ~70 cM away from SCA9-2, suggesting that independent assortment of these QTLs is possible. A digenic epistatic interaction was found between the two main effect QTL on LG A9; and the epistasis × environment interaction was nonsignificant, suggesting stability of the interaction across the environments. The QTL effect on LG A9 was validated using simple sequence repeat (SSR) markers from the two QTL regions of this LG on a B(1)S(1) population (F(1) backcrossed to Sampad followed by self-pollination) segregating for brown and yellow seed color, and on their self-pollinated progenies (B(1)S(2)). The SSR markers from the QTL region SCA9-2 showed a stronger linkage association with seed color as compared with the marker from SCA9-1. This suggests that the QTL SCA9-2 is the major determinant of seed color in the A genome of B. rapa.