Using InDel And SSR Molecular Markers To Integrate A Genetic Map Of Chinese Cabbage (B. Rapa Ssp. Pekinensis)

Brassica rapa crops are cultivated in China. Chinese cabbage(Brassica rapa L.ssp pekinesis) is an important crop which is economical for agriculture and it is a model plant for studies and construction of a molecular genetic linkage map. In2003, it was launched that the multinational B. rapa Genome Sequencing Project (BrGSP) were established. In recent years, some jounals have published that several maps concerning B. rapa genomic sequencing, however, their coverage of the genome is incomplete, and about73.6%of the scaffolds were anchored on to chromosomes. Anyway, a novel molecular genetic map has important research significance.The construction of a genetic linkage map lay the foundation of QTL analysis and map based agronomic trait, molecular marker assisted selection(MAS), functional genes cloning and comparative genomics.Sequence-based insertion/deletion (InDel) markers and SSR markers were used to optimized reasonably and integrated a Chinese cabbage (B. rapa ssp. pekinensis) genetic linkage map. The purpose of the study is to improve the coverage of the genome and identify a specific marker’s position in linkage groups.One hundred and eighty-three doubled haploid (DH) lines were used to construct the linkage map.They were derived from microspore cultures of an Fl cross between a Chinese cabbage (B. rapa ssp. pekinensis) DH line (Y177-12) and line (Y195-93) which were different from each other distinctly in kernelseedcoatcolour, foliar pastel, flower color, bolting time phenotype,disease resistance, et al.In order to ensure experiments proceed smoothly, seven factors (template DNA, annealing temperature, dNTP, et al. for PCR reaction) affecting PCR reaction system were optimized with single factor experiment by making use of DH line parents(Y177-12and Y195-93). Concernning about the stability and economy, the electrophoresis results showed clear bands, which was applied to Brassica rapa PCR amplification conditions of analysis. The optimal amplification conditions(20uL) are template DNA105ng, Taq DNA polymerase0.4U, Mg2+1.5mmol/L,dNTP0.lmmol/L, primer0.4μmol/L, three temperature zones30s-30s-45s, annealing temperature50℃. It can be used in genetic diversity analysis and genetic map construction in Chinese cabbage.They were selected as the most informative ones from401InDels and SSRs between the two parental lines. These selected produced44polymorphic InDels and11polymorphic SSRs. These selected produced55polymorphic markers were integrated a B. rapa genetic linkage map which has SRAP,SSR,AFLP markers in our lab(YX. Yuan,et al.2010). Software JoinMap4.0was used for B. rapa genetic linkage map construction.The map comprises a total of507markers including139SRAPs,84SSRs,44InDels,230AFLPs,6STSs,4ESTPs,3morphological markers,l SCAR and1CAPS. This map has ten linkage groups, designated A0l-A10.The total length of the linkage map was1336cM, with an average distance of2.77cM between adjacent marker loci.The number of markers of linkage groups ranged from27to76on every linkage group.The lengths of linkage groups ranged from106cM to176cM corresponding to2.08-4.41cM per marker on each group. The ratio of Segregation deistortion markers is71.3%.Compared with the previously available molecular linkage maps,the total length, average distance among markers and numbers of markers,this indicators,can be associated with internationally agreed reference linkage map in Chinese cabbage. The development of this linkage map is important for the integration of genetic information, and provides a very useful resource for the international Brassica research.