Charakterisierung genomischer Regionen, die für das außergewöhnliche Mikrosporen-Embryogenese-Potential der doppelhaploiden Linie DH4079 (Brassica napus L.) verantwortlich sind
Zusammenfassung der Projektergebnisse
Microspore culture is a very powerful technique in breeding of oilseed rape for the rapid and efficient generation of completely homozygous doubled haploid (DH) lines. Despite the progress achieved in optimizing tissue culture protocols, tremendous differences remain among rapeseed genotypes in their embryogenic response and direct embryo to plant conversion which are a hindrance in the development of DH populations. The genetic basis of those differences is still largely unknown. To add to our understanding of microspore embryogenesis, a DH population was generated from the cross between the highly embryogenic spring oilseed rape line DH4079 and the low embryogenic winter oilseed rape cultivar Express (Inbred line #617). Replicated experiments were performed for up 97 DH lines out of the complete population of 207 genotypes. Analysis of variance revealed significant genotypic effects for the traits microspore embryogenic potential, survival of embryos, direct embryo to plant conversion, root regeneration and secondary embryogenesis and heritabilities ranged from 66% to 86%. The population of 207 DH lines was genotypes using the Illumina 60K Brassica Chip. Based on 22714 polymorphic markers a frame work map consisting of 1414 SNP markers distributed over 19 linkage groups was developed for QTL mapping. Significantly distorted marker segregation was observed for 48.9% of the SNP markers from which 63.4% showed a significantly higher deviation in favor of the microspore embryogenic responsive cultivar DH4079. Altogether 13 QTL on linkage groups A01, A02, A05, A10, C04 and C06 were identified for aforementioned traits. BLAST-analysis revealed the presence of nineteen candidate genes within the 95 % confidence interval of QTL for microspore embryogenic potential, direct embryo to plant conversion and secondary embryogenesis. Among those, CLE1, CLE5, WOX1, WOX2, WOX5, CUC3, NAM, SCR and SCL11 are putative candidate genes because of their key activity in meristem initiation, development and maintenance. The GA20ox1 gene is a significant candidate for its contribution to seed germination that is associated with direct embryo to plant conversion under cold conditions. A cold treatment of microspore derived embryos has been shown to significantly increase their direct embryo to plant conversion. A MACE analysis (Massive analysis of cDNA ends) performed with cold-treated and control embryos of the two parental genotypes revealed a range of genes whose expression was most significantly up- and downregulated in response to the cold treatment.
