Clonging And Developming STS Markers Of Gene Associated With Drought Resistance In Common Wheat

Wheat is the second largest food crops of China. Drought is one of the important limitingfactors in wheat production. It is one of the effective ways to promote the sustained and stabledevelopment of wheat production and ensure national food security for breeding andpromotion of drought-resistant varieties of wheat. It is very important for breeding wheatcultivars with resistant drought to excavate drought resistance genes and develop theirmolecular marks.In this study, wheat variety jinmai47was used as test material. The full-length cDNA ofa novel gene (TNRX) of thioredoxin (Trx) superfamily was isolated from common wheat bygene homology cloning, electronic cloning, RACE technique and bioinformatics softwareanalysis. Molecular markers of the TNRX gene were developed after genomic sequencevariances between cultivars with higher relative germination rate (RGer) and those with lowerrelative germination rate were analyzed. Those molecular markers were verificatied by150wheat cultivars (advanced lines) and a population from the cross between Jinmai47andXinong2208.The main results obtained in this study are summarized below.1. The RGer of150wheat cultivars (advanced lines) were visible difference. Thedrought resistances of those cultivars were significant differences between the promotion orbreeding ecology of the areas. In those material,15highly resistant and53resistant cultivars(advanced lines) were identified. The RGer and drought resistance index (DRI) of thepopulations from the cross between Jinmai47and Xinong2208were both visible difference.The RGers showed very significant positive correlation (r=0.47, P<0.01) with those DRIsafter Correlation analysised.2. The full-length cDNA of gene TNRX contains1734-bp ORF,(GenBank accessionnumber KC890769). The ORF encodes a protein of577amino acid residues with a putativemolecular mass of63.79kD which may be a liposoluble stabilizing protein, composed ofthree TRX-like modules. The dicots and the monocots were respectively clustering to the twosubsets with blast and phylogenetic analysis the protein encoded by TNRX of wheat and otherhomologous sequence from different species. TNRX gene had the closest genetic relationshipwith the homologous gene of barley and farthest genetic relationship with rice. 3. Gene TNRX has four exons and three introns, mapped on chromosome5BS. Thevariations of TNRX sequences between wheat cultivars Xinong2208and Jinmai47weremainly concentrated in the first intron after genomic sequence variances analysis betweenthem, containing43single nucleotide polymorphisms (SNP),4inserts (Ins) and6deletions(Del). It showed that the sequences of the cultivars with higher relative germination rate(RGer) were same as Jinmai47in the first intron region and the sequences of the cultivarswith with lower RGer were no difference with Xinong2208in the same region.4. Based on the variations of a3bp Del sites and in the immediate vicinity of the threeSNP sites within the first intron of gene TNRX, two markers Tnrxa-1and Tnrxb-1weredesigned. Based on the variations of a28bp Ins sites within the first intron of gene TNRX, twomarkers Tnrxa-2and Tnrxb-2were designed. Two allelic variants TNrx-a and TNrx-b werefound in Triticum aestivum. Marks Tnrxa-1and Tnrxa-2was used to specifically amplify the841-bp and870-bp product from TNrx-a genotypes while had no product from TNrx-bgenotypes. Marks Tnrxb-1and Tnrxb-2was used to specifically amplify the845-bp and910-bp product from TNrx-b genotypes while had no product from TNrx-a genotype. FourSTS markers were complementary dominant markers which could verify each other. Theresult indicated that the variations of the region sequence those development marks relatedhaving synchronization.5. The average RGer with TNrx-a genotype identified by STS molecular markers washigher than those with TNrx-b genotype (P<0.01). It showed that those molecular markerscould be used to identify wheat cultivars with drought resistance and this gene TNRX plays animportant role in wheat tolerance to drought stress.In this study, a number of strong drought-resistant wheat materials were identified. Anovel gene TNRX that mapped on chromosome5BS was first isolated from common wheat.Two allelic variants related drought resistance TNrx-a and TNrx-b were found based on theanalysis of the characteristic of the gene structure and encoding protein. Four complementarydominant STS markers were developed after the genomic sequence variances betweencultivars with higher RGer and those with lower relative germination rate were analyzed. Allthese results provided reference molecular marks for breeding wheat cultivars with resistantdrought and also provided a theoretical basis for further study of wheat drought resistancemechanism.