Construction of a cDNA Libray of the Bovine Viral Diarrhea Virus Genome

MI Ke-Yong; WANG Qiang; ZHU Yong; REN Xiang-Yuan; YANG Yu-Ying

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April 1995

MI Ke-Yong, WANG Qiang, ZHU Yong, REN Xiang-Yuan and YANG Yu-Ying. Construction of a cDNA Libray of the Bovine Viral Diarrhea Virus Genome[J]. Virologica Sinica, 1995, 10(2).

Citation: MI Ke-Yong, WANG Qiang, ZHU Yong, REN Xiang-Yuan, YANG Yu-Ying. Construction of a cDNA Libray of the Bovine Viral Diarrhea Virus Genome .VIROLOGICA SINICA, 1995, 10(2) : 161.

牛病毒性腹泻病毒基因组cDNA文库的构建

1.
中国科学院武汉病毒研究所
2.
内蒙古畜牧科学院

摘要

以牛病毒性腹泻病毒（ＢＶＤＶ）ＩｎＬ株的基因组ＲＮＡ为模板，经逆向转录酶作用，合成第一链ｃＤＮＡ，再以ＲＮａｓｅ／Ｈ与ＤＮＡ聚合酶Ｉ联合作用合成ｄｓｃＤＮＡ，并以ｄＣ同聚物尾化。ｐＵＣ８ＤＮＡ在ＰｓｔＩ酶解后，以ｄＧ同聚物尾化，两者退火构成重组质粒，转化到Ｅ．ｏｏｌｉＪＭ１０１受体菌中。另以γ－￣（３２）Ｐ－ＡＴＰ标记ＢＶＤＶＲＮＡ制备探针，通过菌落原位杂交筛选重组子。酶切分析表明重组质粒插入片段最长接近１２ｋｂ。该片段含有ＥｃｏＲⅠ，ＨｉｎｄⅢ、ＢａｍＨⅠ的酶切位点，此ｃＤＮＡ文库基本上包括了ＢＶＤＶ基因组。
- 牛病毒性腹泻病毒
- , 逆转录
- , 基因克隆
- , cDNA文库

Construction of a cDNA Libray of the Bovine Viral Diarrhea Virus Genome

1.
Wuhan Institute of Virology
2.
Academia Sinica

Abstract

ovine viral diarrhea virus(BVDV)InL genomic RNA was used as a template to synthesizeDNA by reverse transcriptase.The ds cDNA was synthesized by RNase H-DNA polymerase,and waslsetrted into the Pst I site of the pUC8 vector by dC-dG homopolymeric tailing and ligation.After beig transformed to E.coli JM101,the recombinants were screened by in situ hybridization with γ-￣(32)P-ATP labeled purified BVDV RNA,Restrict ion enzyme ana lysis sho wed that the largest cDNA insertwas 12kb-13kb,it almust covered the BVDV...
- BVDV Reverse transcription
- , Gene cloning
- , cDNA library

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Construction of a cDNA Libray of the Bovine Viral Diarrhea Virus Genome

1. Wuhan Institute of Virology
2. Academia Sinica

Abstract: ovine viral diarrhea virus(BVDV)InL genomic RNA was used as a template to synthesizeDNA by reverse transcriptase.The ds cDNA was synthesized by RNase H-DNA polymerase,and waslsetrted into the Pst I site of the pUC8 vector by dC-dG homopolymeric tailing and ligation.After beig transformed to E.coli JM101,the recombinants were screened by in situ hybridization with γ-￣(32)P-ATP labeled purified BVDV RNA,Restrict ion enzyme ana lysis sho wed that the largest cDNA insertwas 12kb-13kb,it almust covered the BVDV...