Expression Analysis Of Three Genes From SSH Library Constructed Using Tillering Nodes Of Dongnongdongmai-1 Under Low Temperature

Wheat as one kind of important crops always is impacted by the environment stress,one of which is the cold-stress,and a lot of reports about the reduction of wheat were publishedd.It is very important to study the molecular mechanisms of stress-resistance and the responses in overall level of molecular expression,and which can provide significant important information to understand the basic knowledge of stress biology and to culture novel species with strong cold resistance. We choose Dongnongdongmai-1(refered as DN-1) as materials to construct the cold-resistant library of tillering nodes through SSH,and then picked 300 white colonies randomly to sequence them, obtained 230 ESTs,of which the shortest was 103bp,the longest was 1215bp,and the average length was 355bp.Apply BLASTx and tBLASTx methods to learn the information of these sequences, 211 ESTs were already-known genes which accounted 91.2% in all genes.Among the 109 genes after BLASTx ,the function of 43 kinds of genes were already known,accounted 18.8% .The homologous putative protein sequences were mainly from barley,wheat,rice, Arabidopsis thaliana,maize.After analysis of ESTs ,we concluded various genes involved in the pathway of cold resistance of DN-1,but the exact mechanism of cold-resistance remains unclear.Among the ESTs, Cold shock protein CS66,cystatin,Clp molecular chaperones,G-protein binding protein maybe participate in the related process of cold-resistance.In addition, there exists many genes involving other biological process, the cold-resistance involved many pathways to realize the function simultaneously .And then examined the three genes in SSH library through real-time PCR,to compare the expression pattern of genes in DN-1 with strong cold-resistance and JM-22 with relative weak cold-resistance,the expression of three genes showed up-regulation in both DN-1 and JM-22 under lower temprature,while the expression in DN-1 showed more robust than JM-22.Therefore, we infered these three genes maybe play an important role in protecting DN-1 enduring the cold winter,and this study can provide a basic beginning for the study of cold-resistance of DN-1.