Development And Assisted Selection Of SNP Markers For Wild Soybean Resistance To Soybean Cyst Nematode

Soybean is one of the main economic crops.Soybean cyst nematode(SCN)is adamaging pathogen of soybean that limits soybean production worldwide.Planting resistant varieties is considered the most effective means against SCN.In the long-term role of natural selection,wild soybeans under different environmental conditions show a strong abilities to adapt,such as high resistance to biotic and high tolerance to abiotic,abundant protein,etc,and provide additional genes for resistant varieties development.Single nucleotide polymorphisms(SNP),as the third generation of DNA molecular markers,can efficiently select wild soybean genetic resources,provide an efficient technique of molecular marker-assisted selection,and accelerate breeding progress inresistant varieties development.QTLs mapping of wild soybean resistance to soybean cyst nematode have been conducting by specific length amplified fragment sequencing,and several QTL loci were identified.In this study,we developed 6 SNP markers basing on information of QTL loci and evaluated their accuracy,analysed their association with resistance,and investigated their effiency in marker-assisted selection.The results were as the following:1.Six SNP markers were developed.We searched for sequence database against the sequence including target SNP locus in Phytomoze v10.3.,and designed allele specific PCR primers for target SNP locus.We successfully got 6 sets of specific primers with their mismatch positions in the 2,3 or 4 position at the end of 3,extremity,and the bases followed the principle of weak or strong in the match.Size range of PCR products for the six SNP markers was 200-400 bp,SNP alleles could be identified by agarose gel electrophoresis.2.Accuracy of SNP markers was evaluated.We employed SNP marker to identify each SNP allele,meanwhile we designed sequencing primes,and identified SNPs allele by sequencing PCR products.We investigated their consistency for each SNP locus to evaluate the accuracy of each SNP marker.The results of five groups specific primers were in accordance with the sequencing results,indicating their accuracy in SNP genotyping.3.Association of SNP markers with SCN resistance were analysed.We genotyped 51 wild soybean accessions using five SNP markers,and evaluated the relationship between SCN resistance and SNP markers.We found that two SNP markers of IP1-1 and IP2-1 were associated with SCN resistance,and the other three of IP5-1,G1-1,and AVE3 were not.4.Efficiency of SNP markers in marker-assisted selection was evaluated.Thirty-three F2 plants from the cross of Suinong14/ZYD03685 were used for efficiency evaluation in marker-assisted selection of two SNP markers(IP1-1 and IP2-1)associated with SCN resistance.We found that the efficiency of two SNP markers in marker-assisted selection were over 65%.So the two SNP markers we have got could be used to screen breeding lineson a large scale rather than phenotyping these lines,because that genotyping using SNP marker is faster and need less labor compared with SCN resistance bioassay.