| 犬瘟热病毒H基因的序列分析与表达 |
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| 引用本文： | 刘刚,ZHANG Yan-ming,李肖梁,WEI Wei,帅江冰,FANG Wei-huan.犬瘟热病毒H基因的序列分析与表达[J].畜牧与兽医,2008,40(8). |
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| 作者姓名： | 刘刚 ZHANG Yan-ming 李肖梁 WEI Wei 帅江冰 FANG Wei-huan |
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| 作者单位： | 1. 西北农林科技大学动物医学院,陕西,杨凌,712100;浙江大学动物预防医学研究所,浙江,杭州,310029
2. 浙江大学动物预防医学研究所,浙江,杭州,310029 |
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| 基金项目： | 浙江省重大科技专项项目 |
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| 摘 要： | 以临床疑似犬瘟热病犬外周血总RNA为模板,采用RT-PCR方法扩增犬瘟热病毒(CDV)血凝素(H)基因。序列测定和分析表明该病料犬瘟热病毒H基因序列与国内其他地区分离到的毒株同源性较高(94.6%~99.2%),而与Onderstepoort株、Convac株等疫苗毒株的同源性较低(90.4%~91.2%)。分子遗传进化分析显示所有毒株可分为7个大的分支,且各分支间具有一定的地域性,其中待检病料中CDV与大多数中国分离株一样处于Ⅰ型。进一步以H全基因为模板,截短表达其3′端816 bp、459 bp两个片段,并克隆入原核表达载体pET30 a进行表达。结果显示它们分别能表达大小约为36.4 ku和22.2 ku的融合蛋白。免疫转印表明该纯化蛋白均可与犬瘟热病毒抗血清发生阳性反应,说明重组蛋白具备抗原性,可用于进一步的血清学研究。
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| 关 键 词： | 犬瘟热病毒 H基因 遗传进化分析 克隆表达 免疫转印 |
Sequence analysis and expression of haemagglutinin(H)gene of canine distemper virus |
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| LIU Gang,ZHANG Yan-ming,LI Xiao-liang,WEI Wei,SHUAI Jiang-bin,FANG Wei-huan.Sequence analysis and expression of haemagglutinin(H)gene of canine distemper virus[J].Animal Husbandry & Veterinary Medicine,2008,40(8). |
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| Authors: | LIU Gang ZHANG Yan-ming LI Xiao-liang WEI Wei SHUAI Jiang-bin FANG Wei-huan |
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| Abstract: | The haemagglutinin(H) gene of canine distemper virus(CDV) was amplified from the blood of a diseased dog by RT-PCR.Sequence analysis showed that the H gene had an identity of 94.6%～99.2% with other Chinese isolates,but only 90.4%～91.2% homology with vaccine strains such as Onderstepoort strain and Convac strain.Phylogenetic analysis revealed that all the CDV isolates were segregated into seven groups which showed some relationships with regard to geographic regions.Obviously,our isolate was clustered in group I together with the most Chinese isolates.Then two fragments of H gene,containing 816 bp and 459bp of its 3′ end,were amplified and cloned into the vector pET30a for prokaryotic expression.The result of SDS-PAGE showed the two bands of 36.4 ku and 22.2 ku as expected.Moreover,the two fusion proteins could be recognized by CDV positive serum in Western blotting,indicating that the proteins had the antigenitic epitopes of H gene of CDV and could be used in serological test. |
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| Keywords: | CDV haemagglutinin gene phylogenetic analysis cloning expression Western blotting |
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