Gene Disruption And Characterization Of BC1G₀4502.1 In Botrytis Cinerea

There 6 genes had been identified involved in transporting monocarboxylic acids in botrytis cinerea.including BC1G₀4502.1?BC1G₁5016.1?BC1G₀0784.1?BC1G₀8724.1?BC1G₀5660.1.In this study,these genes had been analyesd by RT-PCR at first,we found BC1G₀4502.1 had high expression.and a blast research also indicate gene BC1G₀4502.1 in the B05.10 which was one of the subspecies in the bortrytis cinerea was very closely to CCD₅6622.1 in T4.Therefore,it means gene BC1G₀4502.1 was conservative and may have some significant functions.Bioinformatical methods was used to predict the physical-chemical characteristics,hydropathy and hydrophobicity,signal peptide,subcellular localization,transmembrane region,structure and function of BC1G₀4502.1 was,construct protein phylogenetic tree and analyze the key conserved sequence region.The results showed BC1G₀4502.1 was a Transmembrane protein and might was involed in transporting monocarboxylic acid.In this study,the function of BC1G₀4502.1gene was anlyzed by gene deletion at First,an entry vector pETHG-UP-DOWN was constructed by the Gateway system,and the deletion binary vector pCAMBIA-Bar-?03 was generated by LR recombination reaction between pETHG-UP-DOWN and the BC1G₀4502.1 destination vector pCAMBIA-Bar-RfA.Through Agrobacterium tumefaciens mediated transformation,hygromycin-resistant transformants were obtained from Botrytis cinerea B05.10.These transformants were analysed by molecular analysis,and 2 deletion mutants??2d-1??2d-2?were confirmed.And the BC-1 strain was by-product.In the gene-function-testing expriments,mutants growed slowlier than The wild-type strain and ectopic transformant at about 50% rate on the minimal medium plates which just used monocarboxylic acids as the sole carbon source,but they showed the same growth rate when they inoculated on the dicarboxylic acids plates.Mutants also had the same growth rate compared to wild-type strain and ectopic transformant BC-1 on the PDA plates.after 14 days,the mutant stains produced little sclertion,at the same time,wild-type strains and ectopic strains produced much more.Pathogenicity tests on apple and tomato fruits showed no difference almost between wild-type and mutants soon,that means gene BC1G₀4502.1may not have relationship with the virulence of botrytis cinerea.