摘要
应用RT2PCR扩增出番鸭呼肠孤病毒MW9710株的σC基因,序列分析表明σC基因核苷酸数为810bp,编码269个氨基酸,相对分子量为29.4ku,DNAstar软件分析MW9710株的σC核苷酸序列和法国番鸭呼肠孤病毒89026株有93%的同源性,而与禽呼肠孤病毒的同源性仅在21%～25%之间,表明番鸭呼肠孤病毒是一类不同于禽呼肠孤病毒的新病毒。将σC基因片段亚克隆到原核表达载体pET32a中,经IPTG诱导后在大肠杆菌BL21(DE3)中得到高效表达,Western2blot进一步验证了表达产物与番鸭呼肠孤病毒阳性血清反应,表明σC基因表达产物具有免疫原性,为下一步研制番鸭呼肠孤病毒诊断试剂盒和基因工程疫苗奠定基础。
Minor core protein σC gene of MDRV MW9710 strain was amplified by RT-PCR with the primers designed by the reported muscovy duck reoviruses. The PCR products were cloned into pMD18-T vector and identified by enzyme cutting analysis. The sequence results showed that σC gene is 810 bp. It contains one open reading frame between 280-1089 that encodes a protein of 269 amino acids with a molecular mass of 29.4 ku. Nucleotide analysis by the DNASTAR(6.0) software showed that MDRV MW9710 σC is highly related to the MDRV 89026, with 93.0% homology, but only shared 21%～25% homology to the avian reoviruses. It indicates MDRV is possible to be a new revirus that different from ARV.The DNA fragment of σC was subcloned into prokaryotic expression vector pET32a and the specific fusion protein with molecular weight 50 ku was expressed. Western blotting assay indicated that the polyclonal antibody against DRV could recognize the recombinat σC protein. It also provided technical support for developing an advanced gene engineering vaccine against muscovy duck reovirus.
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2005年第4期376-380,共5页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
哈尔滨兽医研究所所长基金(B1-02)
