Genetic And Physical Mapping Of Spot Blotch Resistance Gene Sb3 In Wheat

Bread wheat is one of the most important food crops in the world and has a special significance to human being.However,wheat diseases are major constraint for large yield losses every year and are the major threads in world hunger.Among the wheat diseases,spot blotch caused by Bipolaris sorokiniana,is one of the most important foliar diseases also causing root rot and black embryo in wheat grain which seriously reduced the yield and processing quality.With rising global temperatures and the change in cropping system,the threat of this disease is increasing.Breeding resistant cultivars is the most economical and environmentally safe method to prevent the epidemic outbreaks of wheat disease.Discovering and cloning wheat disease resistance genes are the basis for studying resistance mechanism,host-pathogen interaction,allelic variation and evolution of wheat resistance gene,as well as molecular marker assisted selection in wheat breeding program.In present study,the pathogenic fungal species from the spot blotch diseased leaf samples of wheat line 621-18-3 were isolated,and characterized through morphological and molecular identification identification,and pathogenicity test.Combining comparative genomics analyses,bulked segregant analysis and RNA-Seq(BSR-Seq),and using the available Aegilops tauschii physical map and draft sequences,Chinese spring 3B reference sequence,Hope 3B sequence and the wild emmer TZ-2 BAC library,the fine physical and genetic linkage maps of spot blotch resistance gene Sb3,derived from wheat line 621-7-1,was constructed.The main results are as following:1.Using tissue isolation method,only one fungal species Bipolaris sorokiniana from the diseased leaf samples of wheat line 621-18-3 was characterized,and determined to be the floral diseases(spot blotch)pathogen.2.The wheat line 621-7-1 is highly resistant and 621-18-3 is highly susceptible to B.sorokiniana.Genetic analysis indicated that the spot blotch resistance of 621-7-1 is controlled by a single dominant gene,designated Sb3.Bulked segregant analysis(BSA),simple sequence repeat(SSR)and expressed sequence tag(EST)mapping showed that Sb3 is located on chromosome arm 3BS linked with EST markers SSR markers Xbarc133,Xbarc147 and Xcfp marker Xcfp30.The genetic distance between BE398268 and Sb3 is 1.92 cM,the genetic distance between Xbarcl33,Xbarc147,Xcfp30 and Sb3 is 0.86 cM.3.By applying bulked segregant analysis and RNA-Seq(BSR-Seq),two polymorphic markers closly liked to Sb3 were developed and the spot blotch resistant gene Sb3 was mapped between markers XWGGC5911 and XWGGC4320 in a genetic interval of 2.15 cM.The orthologous genomic region of Sb3 on Aegilops tauschii was identified and the SNP markers extended sequences of the corresponding region from AT3D2320 to AT3D2344 were used to search homoeologous 3BS contigs of the hexaploid wheat cv.Chinese Spring 454 contigs and IWGSC survey sequences for marker development.Seven polymorphic markers,XWGGC6125,XWGGC6135,XWGGC5969,XWGGC6137,XWGGC5963,XWGGC3957 and XWGGC611 9 were developed and the spot blotch resistant gene Sb3 was mapped between markers XWGGC5911 and XWGGC4320 in a genetic interval of 2.15 cM,and co-segregated with marker XWGGC3957.Both polymorphic bands of the markers XWGGC5969 and XWGGC6119 could be mapped on the distal 3BS bin 0.78-1.00 of Chinese Spring deletion lines,indicating that the spot blotch resistance locus Sb3 was also located in 3BS bin 0.78-1.00.4.Using the Chinese Spring 3B reference sequence,fine genetic and physical maps of spot blotch resistance genes Sb3 were constructed.The Sb3 gene was further delimited into a 380 kb genomic region between markers XWGGC9893 and XWGGC12530 in Chinese Spring,and is corresponding to physical distance of 708 Kb in wheat cultivar Hope.Molecular markers tightly linked to Sb3 were used as probes to screen a Hind? BAC library of wild emmer accession TZ-2 to construct a partial physical map of Sb3 in T.dicoccoides.Comparative analysis between the Chinese spring,Hope and TZ-2 revealed two resistance-related genes,one receptor-like kinase(RK)TaRK and one Ser/Thr receptor-like kinase(STK)TaSTK,could be served as candidates for Sb3.