猪源冠状病毒ORF3和ORF7的克隆及特性分析

参照GenBank中Purdue株全序列对TGEV(猪传染性胃肠炎病毒)TS株的ORF3和ORF7各设计1对特异性引物,经RT-PCR扩增分别获得了1 487 bp和516 bp大小的两个片段,与预期结果大小相符.TS株与CHV株、Puedue株、BW021898B株和KT2株的ORF3a核苷酸序列同源性分别为99.1%、93.0%、95.9%、94.4%,相应的氨基酸同源性分别为97.3%、87.8%、93.2%、91.8%,TS株与CHV株、Puedue株、BW021898B株和KT2株的ORF3b核苷酸序列同源性分别为99.5%、98.5%、97.0%、97.8%,相应氨基酸的同源性分别为98.4%、96.3%、94.3%、95.8%,而TS株与PRCVIA1894株的ORF3b核苷酸和氨基酸同源性分别为96.6%和93.9%.TS株ORF7没有发生缺失且和其他毒株有很高的同源性.结果表明:TGEV非结构蛋白高变区在ORF3a第192个碱基至终止密码子之间,TGEV和PRCV在编码ORF3b氨基酸的数量和RNA酶结合位点上有一定的差异.

Cloning and characterization of ORF3 and ORF7 of porcine coronavirus

Two non-structural protein genes of strain TS of transmissible gastroenteritis virus(TGEV),ORF3 and ORF7 were obtained by RT-PCR using primers designed according to Purdue nucleotide sequence in the GenBank,with the length of 1 487 bp and 516 bp respectively.The homology at the nucleotide levels for the ORF3a gene between TS and CHV,Purdue,BW021898B,KT2 was found to be 99.1 %,93.0 %,95.9 % and 94.4 % respectively,and that of the deduced amino acid sequence was 97.3 %,87.8 %,93.2 %and 91.8 % correspondingly.The nucleotide homology of ORF3b gene of TS shared 99.5 %,98.5 %,97.0 % and 97.8 % nucleotide identity and 98.4 %,96.3 %,94.3 % and 95.8 % deduced amino acid with that of strains CHV,Purdue,BW021898B,KT2.The results indicated that there was no deletion in the TS ORF7 and it was conserved.The region from the 192 to TAA in the ORF3a was considered as the region of high variety of TGEV,the results showed that there was differences in the num ber of amino acid and the RNA-binding site between the TGEV and PRCV.