关键词: 马铃薯卷叶病毒, CP 基因, 缺失突变, 原核表达, 抗血清

Abstract: Potato leafroll virus（PLRV）coat protein（CP）gene（PLRV-CP）was amplified by RT-PCR，and the specific product about 630 bp was cloned through T-A ligation. DNA sequencing showed that its length was 627 bp，and the homology of nucleotide sequences between the cloned PLRV-CP gene and other 36 previously published ones was above 96%. Using pBAD/Thio-TOPO as initial vector，prokaryotic expression vector of PLRV-CP gene was constructed and named pBAD-LRCP. DNA fragment（52–177 nucleotides）rich in rare codons of arginine was deleted from PLRV-CP gene by PCR using pBAD-LRCP as template，the yielding recombinant plasmid pBAD-LRCP-126 was indeed the prokaryotic expression vector of deletion mutation CP gene of PLRV. After induction of the engineered strain TOP10（pBAD-LRCP-126）with arabinose，the mutation gene was successfully expressed and 34 kD fusion protein（the recombinant CP）was obtained. High-purity recombinant CP was purified from inclusion bodies with nickel affinity chromatography column，and the purified recombinant CP was used as antigen to immune rabbits. Antiserum specific to PLRV was obtained，its titer was 1︰12 800 in indirect ELISA detection. This research has laid the foundation for preparation of antiserum against PLRV by using recombinant CP as antigen in a large scale.

Key words: Potato leafroll virus, CP gene, deletion mutation, prokaryotic expression, antiserum