柑橘CsERF1基因RNAi载体的构建及转化

ERF1(ethylene response factor 1)is a intranuclear transcription factor involved in the end of the ethylene signal transduction pathway.The sense and anti-sense cDNA sequences of CsERF1 were amplified from Citrus by RT–PCR and cloned into pMD–19T respectively,resulting in plasmids containing sense and anti-sense fragment of CsERF1 respectively.Plasmid containing sense CsERF1 and pFGC5941 were digested by XhoI and AscI,and the sense fragment of CsERF1 was inserted into binary vector pFGC5941 forming+CsERF1–pFGC5941,then the anti-sense fragment of CsERF1 was subsequently cloned into+CsERF1–pFGC5941 by SmaI and XbaI digestion and ligation of +CsERF1–pFGC5941 and plasmid containing anti-sense CsERF1,resulting in RNAi vector of CsERF1 with sense and antisense fragment flanking the intron of chalcone synthase a gene.Verified by PCR testing and sequencing,the RNAi vector of CsERF1 was successfully constructed.Antibiotic resistant buds were regenerated via Agrobacterium-mediated transformation of citrus explants which provides a way towards the elucidation of the function of CsERF1.