Genetic variation and inheritance of phytosterol and oil content in winter oilseed rape (Brassica napus L.)

Phytosterols are natural constituents of vegetable oils that are among the dietary option to reduce serum LDL-cholesterol. Oilseed rape (Brassica napus L.) is ranked the second richest source of phytosterols among vegetable oils. Improving the phytosterol content or composition could enhance the nutritive value of oil produced from oilseed rape. To understand the genetic basis of oil and phytosterol content and its relationship with other traits, QTL analysis was performed with both linkage mapping and association mapping. Linkage mapping was performed on a double haploid (DH) population derived from the cross of two winter oilseed rape varieties "Sansibar" and "Oase" and was termed SODH population. Both parental lines are of canola quality and were chosen due to their contrasting phytosterol and oil content in seed. A genetic map was constructed for SODH population based on a total of 1642 markers (AFLP, candidate-gene based marker, DArT, Silico-DArT, SSR, and SNP), organized in 23 linkage groups and covering a map length of 2350 cM with a mean marker interval of 2.0 cM. Field trials of SODH population were carried out at two mega-environments, Europe (Germany & Sweden) and East China (Hangzhou).  Association mapping was conducted on 81 canola quality winter oilseed rape varieties and breeding lines with a total of 692 markers (685 AFLP and 7 SNP/InDels candidate gene-based markers) using K model--a mixed linear model (MLM) which incorporates kinship matrix (K) as a random effect. Field trials of association mapping population were performed at six environments in Germany. Total phytosterol content in the SODH population ranged from 311.2 to 486.9 mg 100 g-1seed in the Europe trial and 340.5 to 507.7 mg 100 g-1seed in the East China trial while total phytosterol content in the collection of 81 canola quality varieties and breeding lines ranged from 298.8 to 491.5 mg 100 g-1seed.  Oil content in the SODH population ranged from 41.2 to 48.6% in the Europe trial and 39.9 to 45.8% in the East China trial while the oil content in the collection of 81 canola quality varieties and breeding lines ranged from 39.1 to 46.8%. Estimates of broad-sense heritability (H2) ranged from 0.82 to 0.94 for total phytosterol content and 0.70 to 0.86 for oil content. Highly significant positive correlation between total phytosterol content and oil content was observed in both populations. Linkage mapping identified between 1 and 6 QTL for the nine phytosterol traits in both trials, 6 QTL for oil content in the Europe trial and 7 QTL for oil content in the East China trial. Comparison of QTL identified across the two trials showed that most traits contain between 1 and 4 stable QTL except for total phytosterol content which showed only environment-specific QTL. Association mapping identified between 1 and 7 QTL for five of the nine phytosterol traits and 6 QTL for oil content. Comparison of QTL results between linkage mapping and association mapping showed that five QTL were found detected at similar positions in both populations. By aligning sequence-informative markers to the reference sequence of B. rapa genome, possible candidate genes were identified for three of the five QTL: (1) QTL brassicasterol on A04 collocated with CYP710A1, a gene that encodes the cytochrome P450 enzyme which might be responsible for converting 24-epi-campesterol to brassicasterol; (2) QTL for 24-methyl:24-ethyl sterol on A06 collocated with SMT2, a gene that encodes the sterol C24-methyltransferase 2 which converts 24-methylenelophenol to 24-ethylidene lophenol, and; (3) QTL for oil content on A01 collocated with LPAAT which plays an essential role in the synthesis of phosphatidic acid, a key intermediate in the biosynthesis storage lipids in developing seeds.