猪繁殖与呼吸综合征病毒贵州PT分离株Nsp2基因克隆及序列变异分析

为了解猪繁殖与呼吸综合征病毒(porcine reproductive and respiratory syndrome virus,PRRSV)贵州PT分离株Nsp2基因序列特征,选择PRRSV JXA1株Nsp2基因保守区域设计并合成特异性引物,用RT-PCR方法扩增出PRRSV贵州PT分离株Nsp2基因片段,克隆至pMD18-T载体并测序,应用DNAStar、PSORTⅡ、TMHMM、MLRC等软件对Nsp2序列的理化参数、同源性、亲水性、表面可及性、抗原性和亚细胞定位等进行分析和预测。结果显示,该基因cDNA长588 bp,蛋白理论分子质量为21.5674 ku,理论pI值为5.02,为不稳定蛋白,具有良好的形成抗原表位的条件。亚细胞定位结果表明该蛋白极有可能位于细胞核,其Nsp2基因与所选18个毒株相应片段的同源性为80.0%~99.8%。预测Nsp2蛋白具有良好的抗原性,在PRRSV的流行过程中,Nsp2基因不断发生遗传变异,产生新的变异序列。

Cloning and Sequence Analysis of Nsp2 Gene of Porcine Reproductive and Respiratory Syndrome Virus Guizhou PT Isolate

In order to know the gene sequence characteristics of porcine reproductive and respiratory syndrome virus (PRRSV) Guizhou PT isolate,a pair of specific primers of Nsp2 gene was designed according to the gene sequence of PRRSV JXA1 strain. The specific fragment of PRRSV Guizhou PT isolate was amplified by RT-PCR and sequenced after being cloned directly into pMD18-T. The softwares of DNAStar,PSORTⅡ,TMHMM and MLRC were used to predict the physical and chemical properties,homology,hydrophilicity,surface probability plot,antigenic index,secondary structure,subcellular localization. The results showed that the cDNA of Nsp2 was 588 bp;the predicted molecular weight was 21.5674 ku and pI was 5.02;it was an unstable protein. The study showed that Nsp2 possessed potential antigenicity and it mainly existed in the nucleus. It shared 80.0% to 99.8% of nucleotide homology with the 18 strains logged in GenBank. The conclusion showed that Nsp2 protein might have good antigenic characteristics,and genetic variation of Nsp2 gene was constantly emerging.