Prokaryotic expression and antiserum preparation of coat protein of rosa rugosa leaf distortion virus
Rosa rugosa leaf distortion virus(RrLDV)belongs to the genus Pelarspovirus in the family Tombusviridae,which can infect roses and cause symptoms such as leaf deformity,yellowing and premature aging.In order to establish a rapid detection method for detecting RrLDV,total RNA of susceptible rose leaves was extracted by hot borate method.The gene which encodes coat protein of the virus was amplified by RT-PCR and constructed to the prokaryotic expression vector pDB-His-MBP.The vector was transformed into Escherichia coli BL21 and the expression of the recombinant protein was induced by IPTG.The recombinant protein with a concentration of 8 mg/mL was obtained and the polyclonal anti-rabbit antiserum was produced.Western blotting showed the titer of the antiserum was 1∶2 048 000 and its sensitivity was 1∶1 024 when the antiserum was diluted to 1∶1 000.The serological detection result was consistent with RT-PCR,indicating that the antiserum can be used to detect the occurrence and distribution of RrLDV.
rosa rugosa leaf distortion viruscoat proteinprokaryotic expressionantiserum preparation
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