The Gene Expression In Different Corn-Setosphaeria Turcia Interactions

Northern corn leaf blight caused by Setosphaeria turcica is one of the important corn leaf diseases. Breeding and growing resistant corn species is the most economical and efficient method to control Northern corn leaf blight. Therefore, separation and cloning of the expressed gene fragments related to corn resistance to Setosphaeria turcica, and defmation of identity and function of their products is significant to knowing of molecular mechanism on corn resistance to Setosphaeria turcica and genetic engineering breeding.The expression of gene fragments in the corn leaves were studied both in incompatible combination (B37Ht1 inoculated with race 0 of Setosphaeria turcica) and in compatible combination (B37Ht1 inoculated with race 1 of Setosphaeria turcica) by mRNA differential display PCR (DDRT-PCR). The fragments that were expressed exclusively in incompatible combination were cloned, sequenced and homogenously analyzed.The main results of the study are as follows:①Polymerase chain reaction (PCR) and Polyacrylamide gel electrophoresis (PAGE) were used to examine 26 random primers.The results indicate that 6 of them were suitable for the one-step amplification method, of which annealing temperature is 40~47℃, and 9 of them were suitable for the two-step amplification method, of which annealing temperature of the second amplification is 46~51 ℃.②The random primers picked out were used for DDRT-PCR of cDNA samples. 108 differential expressed gene fragments were obtained by PAGE and silver stain detection.These fragments were divided into three classes in incompatible combination compared with either in compatible combination or in control: A.26 gene fragments were expressed exclusively, and the percentage is 24%;B.39 gene fragments expressed increasingly, and the percentage is 36%;C.42 gene fragments expressed decreasingly, and the percentage is 40%.98 fragments of them were successfully recovered from PAGE gel and were effectively amplified again by Agarose electrophorsis detection,and the recovery percentage was up to 92%.(3) Through reverse Northern blotting, 46 positive fragments were obtained, and the positive percentage is 46.9%. Among them, 10 fragments belonged to A, and the percentage is 21.8%;19 fragments belonged to B, and the percentage is 41.1%;17 fragments belonged to C, and the percentage is 37.1%.(4) Five fragments out of A were cloned,sequenced and homogenously analyzed. The experimental result showed that the identity of them were all low with known protein sequence, which is 30%~40%. To understand function of the fragments, the full-length genes should be obtained and their functional complement experiments should be performed in the next step.