Cloning And Prokaryotic Expression Of Wheat β-1, 3-Glucanase Gene Induced By Bipolaris Sorokinianum

Wheat root rot is a kind of serious disease in wheat worldwide. Breeding resistant varieties is an efficient way in controlling wheat root rot.Therefore, it is very important for breeding resistant varieties to clone resistant-related genes and to confirm their roles in the interaction of wheat and pathogenic.β-1,3-glucanase was an important pathogen-related protein. It could catalyze the hydrolysis ofβ-1,3- glucan that was one of the major components of most pathogenic fungal cell walls, which made cell inclusion overflow and dead at last.Firstly, glu gene was amplified by RT-PCR from wheat leaves inoculated with Bipolaris sorokinianum, and was introduced into pMD18-T vector subsequently. The recombinant clone pMD18-T/glu was sequenced. The nucleotide sequence of glu was compared and analyzed with the one published previously in GenBank, then, pET-30/glu recombinant plasmid was constructed by inserting glu gene into pET-30a vector. The recombinant plasmid was transformed into E. coli BL21 strain, and was induced with IPTG to express recombinant GLU fusion proein. The expression of target protein was detected by SDS-PAGE. Bioinformatics analyze glu gene. The cDNA sequence and character of glu were analyzed in NCBI and EXPASY Server.The results of the study showed that 1 005 bp lengthed glu gene of wheat was successfully amplified and cloned. Sequencing results showed that the glu gene of wheat shares 100% homology in nucleotide sequence of the DQ090946 in GenBank. SDS-PAGE result showed that the recombinant pET-30/glu was expressed successfully in E. coli LB21 strain. Bioinformatics analyze glu gene, GLU is extracellular (secreted) protein.These results suggest thatβ-1,3-glucanase gene can be expressed aftet Bipolaris sorokinianum inoculated.