Development of intervarietal substitution lines in <i>Brassica napus</i> L. using marker assisted selection and mapping of QTL for agronomically important traits

Abstract

Brassica napus L. is widely grown as oil crop in temperate areas of North America, Europe and China. In this study, intervarietal substitution lines were developed and were used for QTL mapping. Substitution lines comprise a set of lines with complementary donor segments in the genetic background of a recurrent parent covering the genome of the donor parent. These lines have a common genetic background except for the introgressed region of the donor segment. The first objective of the current study was the development of two intervarietal substitution lines libraries from crosses of rapeseed varieties. The second objective was to map QTL for agronomically important traits using one of the intervarietal substitution lines libraries.The first library was developed using Express as the recurrent parent and R239 , a resynthesized rapeseed, as the donor parent. A map with 220 AFLP markers covering 1327 cM of the rapeseed genome was used in the development of the substitution lines. After 4 generations of backcrossing to the recurrent parent and marker assisted selection, a total of 27 BC4 plants were used to develop doubled haploid substitution lines using microspore culture. The second substitution line library was developed using Mansholts Hamburger Raps as the donor parent and Samourai as the recurrent parent. 164 AFLP markers that cover 1325 cM of the rapeseed genome were used for marker assisted selection in the development of the substitution lines. A total of 26 genotypes were selected in BC4 and 15 seeds from each plant were sown to produce 390 genotypes. The 390 genotypes were analysed by AFLP markers. A total of 76 genotypes were selected with donor segments covering 525 cM of the mapped rapeseed genome. The 76 selected BC4 plants were used to produce doubled haploid lines with homozygous donor segments using microspore culture.The substitution lines from the cross of Mansholts Hamburger Raps were used for QTL mapping of agronomically important traits. To identify the QTL, multiple comparisons of least square means differences of the substitution lines to the recurrent parent were tested with t-tests adjusted according to Dunnet. A total of 10 QTL were mapped for oil content with QTL effects ranging from -5.7 to 6.2% of which 9 QTL increased oil content in the substitution lines. The donor parent contained lower oleic acid content compared to the recurrent parent. For oleic acid content 4 QTL with QTL effects ranging from -12.3 to 13.7% were identified of which 3 QTL decreased oleic acid content. For protein content a total of 5 QTL were identified with QTL effects ranging from -3.9 to 3.1% and except one all QTL effects showed a decrease in protein content. Four out of five QTL for protein content also affected oil content in the opposite directions of the effects on protein content implying a close linkage of the QTL or pleiotropic gene effects. The reason for the complementary relation could be due to the same substrates being used in the synthesis of oil and protein. However, one QTL for protein content independent of any effects on oil content was also identified.The donor parent is characterized by high glucosinolate content. A total of 9 QTL were identified and all of the QTL effects showed an increase in glucosinolate content ranging from 12 to 28 µmol/g seed due to the replacement of Samourai alleles by alleles of Mansholt . The donor parent survived winter better than the recurrent parent. 13 QTL were detected for winter survival due to the introgression of donor segments. Erucic acid genes were also examined in relation to content of phytosterols and sinapate esters. Erucic acid genes on linkage groups 6 and 12 decreased total sinapate ester content by 1.26 and 0.99 mg sinapate g-1 seed, respectively, compared to erucic acid free lines. Moreover, erucic acid genes on linkage groups 6 and 12 decreased total phytosterol content by 469 and 622 mg kg -1 seeds, respectively, compared to erucic acid free lines.