Cloning and Prokaryotic Expression of NS1 Gene of Porcine Parvovirus (PPV) SD1 Strain

猪细小病毒(PPV)SD1株NS1基因的克隆与原核表达(英文)

[Objective] The research aimed to provide the theoretical basis for establishing a rapid diagnosis method for porcine parvovirus（PPV）. [ Method] One pair of primers were designed according to PPV genome sequences on GenBank website and the sequences of prokaryotic expression vector pET30a （ ＋ ） with multiple cloning sites. The whole sequence of NS1 gene in PPV SD1 strain was amplified by using PCR technology and the positive recombinant plasmid was analyzed by sequencing and homology comparison. The prokaryotic expression recombinant plasmid PET30a/NS1 was constructed to make its induction expression in Escherichia coll. [ Result] The target fragment with the length of 2 208 bp was obtained from PCR amplification. The nucleotide homologies between the cloned NS1 gene and the reported relevant PPV genes were from 97.3 % to 99.4 %, which indicated that NS1 gene had high conservation. But it had a 12-basepair successive deletion near the hydroxyl end. The cloned PPV NS1 gene was successfully expressed in prokaryotic cell, and its expression products existed mostly in inclusion bodies. [ Conclusion] The results of SDS-PAGE detection showed that the molecular weight of PPV NS1 protein was 86 KD.

[Objective] The research aimed to provide the theoretical basis for establishing a rapid diagnosis method for porcine parvovirus(PPV).[Method] One pair of primers were designed according to PPV genome sequences on GenBank website and the sequences of prokaryotic expression vector pET30a(+)with multiple cloning sites.The whole sequence of NS1 gene in PPV SD1 strain was amplified by using PCR technology and the positive recombinant plasmid was analyzed by sequencing and homology comparison.The prokaryotic expression recombinant plasmid PET30a/NS1 was constructed to make its induction expression in Escherichia coli.[Result] The target fragment with the length of 2 208 bp was obtained from PCR amplification.The nucleotide homologies between the cloned NS1 gene and the reported relevant PPV genes were from 97.3 % to 99.4 %,which indicated that NS1 gene had high conservation.But it had a 12-basepair successive deletion near the hydroxyl end.The cloned PPV NS1 gene was successfully expressed in prokaryotic cell,and its expression products existed mostly in inclusion bodies.[Conclusion] The results of SDS-PAGE detection showed that the molecular weight of PPV NS1 protein was 86 KD.