Penicillium Marneffei Rho Gtpase Activation Factor Rna Interference Vector And Its Functions Preliminary Study

Penicillium marneffei is the only known species of its genus that is temperature-dependent dimorphic.Now,it has emerged as one of the most common opportunistic pathogen causing respiratory,skin and systemic mycosis among immunocompromised and AIDS patients,endemic in South-east Asia and Southern China.To obtain a better understanding of the key elements involved in Penicillium marneffei virulence and to identify possible drug targets,it is necessary to be able to study the gene fuction.Unfortunately,the molecular techniques available do not include a rapid method to disrupt and identify essential genes.RNA interference,a process in which the presence of double-stranded RNA homologous to a gene of interest results in specific degradation of the corresponding message,has been known as powerful and useful tools in gene functional research.Rho GTPase may play an important role in the polar growth of cell.We are particularly interested in understanding the function of Rho GTPase using RNAi method instead of the traditional method.The Rho GTPase activator gene including xylp promoter and gus gene was cloned into pCB309A vector with ble~R inserting into T-DNA region.Bleomycin resistant gene was used as selecting marker to select the mutants after Agrobacterium-mediated transformation in Penicillium marneffei.We have shown that expression of double stranded RNA corresponding to portions of the Rho GTPase activator gene results in reduced mRNA levels,with phenotypic consequences similar to that of gene disruption.Because of the important role that Rho GTPase played in the eukaryotes and the establishment of gene silencing transformation system,we do initial research on the fuction of Rho GTPase.The results show that this gene may control morphogenesis in Penicillium marneffei at 25℃.It is the first report of RNAi for Rho GTPase activator in Penicillium marneffei by Agrobacterium-mediated transformation.Use of RNA interference in Penicillium marneffei will allow easier examination of the phenotypic consequences of reducing expression of a gene of interest,especially for essential genes.Also,this study provides a basis for future in-depth nsights in the biology of Penicillium marneffei and its pathogenetic mechanism.