Construction Of An SSH Library From Haynaldia Villosa Leaves Induced By Erysiphe Graminis And Cloning And Analyses Of Three Powdery Mildew Resistance-related Genes

Powdery mildew is one of severest diseases in wheat production. Wild Haynaldia villosa contains Pm21 gene which has wide and strong resistance to Powdery mildew. Clomng of Pm21 gene and Powdery mildew resistance-related genes will play an important role in future disease resistance breeding. In this research, we attempted to construct an SSH (suppression subtractive hybridization) library using the leaves of wild H. villosa (Gh21) inoculated by Erysiphe graminis and its susceptible EMS-induced mutant M14S with no inoculation.We screened 218 positive clones in the SSH library, and gained 60 ESTs after sequencing. Similarity analysis was based on BLASTx and BLASTn softwares in GenBank. Twenty-three of them might be related to powdery mildew resistance, including one resistance gene analog(RGA), two effector in pathogen defense, three proteins related to signal transduction, five enzymes to important secondary metabolism, four enzymes to protein degradation, three proteins or enzymes to repairation or fortification of cell wall or membrane and five reactive oxygen species(ROS) scavengers. Twenty-nine of them involve in response to abiotic stresses, RNAs processing, proteins translation and processing, photosynthesis, energy metabolism and other pathways. Five of them were found similarity in GenBank, but their functions are unclear. Three of them were found no significant similarity and they may represent new genes or high variant non-coding regions of cDNAs.In this research, we screened a blue copper-binding protein (BCB) gene and a glutathione-S-transferase (GST) gene from cDNA library of H. villosa. And we cloned three cyclophilin (CyP) genes by PCR according to wheat CyP sequence retrieved from GenBank. Expression anlyses by semi-quantitative RT-PCR showed that all of these genes were expressed constitutively and their products were accumulated greatly after infected by E. graminis. At the same time after infection, expression level of BCB gene had no significant difference between Gh21 and M14S, and expression level of GST gene was higher in Gh21 than that in M14S. CyP gene had different expression pattern in the two materials. These results revealed their possible different functions in powdery mildew defense.