Molecular tagging of erucic acid trait in oilseed mustard (Brassica juncea) by QTL mapping and single nucleotide polymorphisms in FAE1 gene

Abstract

Molecular mapping and tagging of the erucic acid trait (C22:1) in Brassica juncea was done by a candidate gene approach. Two QTLs underlying the variation of seed erucic acid content were assigned to two linkage groups of a B. juncea map using a doubled haploid (DH) mapping population derived from high x low erucic acid F(1) hybrid. Two consensus primers corresponding to the full-length Fatty Acid Elongase 1 ( FAE1) gene, reported to be involved in the elongation of C18:1 to C22:1, were designed. PCR amplification and subsequent cloning and sequencing identified two FAE1 genes ( FAE1.1 and FAE1.2) in both high and low erucic acid mustard lines. Sequence alignment of corresponding FAE1 genes between high and low erucic acid mustard lines identified four substitution type single nucleotide polymorphisms (SNPs) in FAE1.1 and three in FAE1.2. Using the SNuPE method of SNP genotyping, these two genes were mapped to two independent loci that co-segregated with the two QTLs governing the erucic acid trait. Association of wild ( E1E2) and mutant ( e1e2) haplotypes of two FAE1 genes with erucic acid variation in two segregating populations revealed that the e1e1e2e2 genotype identified low erucic acid individuals (<2%) and E1E1E2E2 identified individuals with highest erucic acid content (>40%). The E1e1E2e2 heterozygote was found to be intermediate in phenotype. The applicability of these SNPs in marker-assisted manipulation of the erucic acid trait was verified by genotyping a set of contrasting germplasm of B. juncea belonging to two distinct gene pools (Indian and east European) and other oil-yielding Brassica species.

Novel Insights into Seed Fatty Acid Synthesis and Modification Pathways from Genetic Diversity and Quantitative Trait Loci Analysis of the Brassica C Genome

“…Qiu et al (2006) reported seven QTL and Ecke et al (1995) reported three QTLs associated with oil content in B. napus. In both studies QTL showed close linkage to the 22:1n9 genes fatty acid elongase1.1 (FAE1.1) and FAE1.2, and a similar situation has been reported for B. juncea (Thormann et al, 1996;Cheung et al, 1998;Gupta et al, 2004). Genetic characterization of the loci involved in 18:1n9 content has been carried out in B. napus, B. rapa (Tanhuanpaa et al, 1998), and B. juncea (Sharma et al, 2002).…”

“…Closely related orthologs of these genes have been isolated from Brassica species. The B. napus genes FAE1.1 and FAE1.2 have been shown to cosegregate with two loci that govern 22:1n9 production (Barret et al, 1998;Fourmann et al, 1998), with FAE1 genes also being isolated from B. oleracea and B. rapa (Das et al, 2002) as well as B. juncea (Gupta et al, 2004). Between four and six copies of FAD2 have been reported for B. napus (Scheffler et al, 1997) and representative orthologous genes encoding for both FAD2 and FAD3 have been cloned (Arondel et al, 1992;Okuley et al, 1994).…”

Intergeneric Somatic Hybridization Between Brassica napus L. and Sinapis alba L.

“…Meanwhile, in the BC 1 progenies, some plants displayed additive patterns from both parents and some had only one specific band from S. alba, demonstrating segregation in the BC 1 progenies. The fae1 gene has been shown to be the candidate gene for elongation of C 18:1 to C 22:1 (Gupta et al 2004). It has been suggested that the variation at the erucic acid level is due to differences in the fae1 gene from the low and high erucic acid genotypes (Das et al 2003).…”

“…The ratio of seed set was measured by the number of seeds obtained per pollinated flower. 1.2.6 Characterization of fae1 gene To clone the full-length fae1 gene from plants of the fusion parents, hybrids, and their progenies, primers were designed according to sequences (Gupta et al 2004). The PCR was performed in a final volume of 20 µL containing 10 ng genomic DNA, 2.5 mmol/L MgCl 2 , 0.2 mmol/L of each dNTP, 0.2 µmol/L of each primer, 1 unit HotStarTaq DNA polymerase and 1×Taq polymerase buffer as follows: 15 min at 95 °C, 35 cycles of 30 s at 94 °C, 30 s at 56 °C, 45 s at 72 °C, and 10 min at 72°C…”

Partitioning genetic effects due to embryo, cytoplasm and maternal parent for oil content in oilseed rape (Brassica napus L.)

“…Recently, quantitative trait locus (QTL) mapping has been conducted on oilseed rape to dissect the effects of individual loci for seed traits in terms of rape-oil yield (Tanhuanpää et al, 1996;Thormann et al, 1996;Zhao et al, 2005). QTL analysis in which the molecular marker near the available locus can be screened is important in molecular breeding strategies such marker associated selection (MAS) used to screen the target materials precisely and improve selection efficiency for quantitative traits (Tanhuanpää et al, 1996;Somers et al, 1998;Gupta et al, 2004). So far, QTL mapping for seed traits has been based on the maternal plant because of the QTL method used to map agriculturally plant traits.…”