Construction Of An SSH Library From Oryza Sativa Leaves Induced By Activator Protein

Plant activator protein is a new type protein separated from many pathogeny fungi of plant. It is different from the Harpin and Elicitin proteins in nucleic and amino acid sequences, and has less identities with proteins in the database of GenBank. Plant activator is a kind of thermostable protein abstracted from fungi, which can promote plant growth, activate the defense system of plant and induce resistance. As a brand-new microbial proteinaceous pesticide, it is high-performance, innoxious, multi-function, broad-spectrum and environment-friendly. The activator protein will have a broad application prospect.In order to understand the molecular mechanism of its function, Suppression subtractive hybridization(SSH) was used in this study to find the different expression in rice reduced by activator protein. SSH is a method to isolate differentially expressed genes, which based on subtractive hybridization and suppression PCR. SSH is widely used in the differentially expressed gene cloning in biology and iatrology due to its high efficiently, stability and reliability.In this study we used activator protein separated from Magnaporthe griesea. A forward suppression subtractive cDNA library of rice leaves regulated by activator was constructed, using the rice treated with activator protein as tester and untreated as driver.The SSH library was totally composed of 1756 recombinant clones. 267 of them were sent for sequencing after Dig detection. Finally we got 204 sequences. After similarity analysis based on BLASTx and BLASTn in GenBank and removed the redundant cDNAs, 28 differentially expressed genes were obtained. 18 genes of them were known or putative proteins. These genes were related to various physiochemical functions of plant. Four of them involved in the construction of chloroplast; Three of them involved in the nutrients substance metabolism; Others were related to other functions such as cells cyclical regulation, intracellular organelle movement, signal transduction, gene regulation, defense response and so on. Ten of them were found similarity sequence in the database, but the functions still not clear. This research established foundation to reveal the molecular mechanism of activator protein.