An efficient method of constructing homologous recom binant baculovirus with pcr-amplified fragments
文献类型：期刊论文
| 作者 | Hou, SW; Chen, XW; Wang, HZ; Hu, ZH |
| 刊名 | Science in china series c-life sciences
 |
| 出版日期 | 2003-08-01 |
| 卷号 | 46期号:4页码:431-437 |
| 关键词 | Phage x Red system Baculovirus bacmid Gene replacement Gene deletion Hasnpv Recombinant virus |
| ISSN号 | 1006-9305 |
| DOI | 10.1360/yc0066 |
| 通讯作者 | Hu, zh() |
| 英文摘要 | This paper describes a rapid method of constructing homologous recombinant baculovirus in e. coli with pcr-amplified fragments. by using this method, the traditional steps of constructing transfer vector are omitted. the method is based on phage lambda, red system which can promote the recombination between the homologous fragments with the length above 36 bp. taking hasnpv as an example, this paper describes the rapid recombination process by using chloramphenicol resistance gene (cm-r) to replace orf135 in hasnpv genome. a pair of primers with length of 60 by was synthesized, in which 40 by was homologous to the each end sequence of orf135, and the rest 20 by was homologous to the each end sequence of cm-r. by using these primers, a linear fragment containing the complete cm-r gene between 40 by of homologous arms of orf135 was generated by pcr with the plasmid pkd3 which contains cm-r as the template. by transforming the linear fragment into the e. coli containing the bacterial artificial chromosome of hasnpv and with the help of a plasmid expressing lambda recombinase, the recombinants on which the homologue replacement had taken place were selected by chloramphenicol resistance. this method greatly shortens the process of constructing recombinant baculovirus since the process was performed in e. coli and does not need to construct transfer vectors. it can be further used for gene replacement and gene deletion of other large viral genomes. |
| WOS关键词 | ESCHERICHIA-COLI ; NUCLEOPOLYHEDROVIRUS ; REPLICATION ; CELLS |
| WOS研究方向 | Life Sciences & Biomedicine - Other Topics |
| WOS类目 | Biology |
| 语种 | 英语 |
| WOS记录号 | WOS:000184455900010 |
| 出版者 | SCIENCE CHINA PRESS |
| URI标识 | http://www.irgrid.ac.cn/handle/1471x/2376136 |
| 专题 | 武汉病毒研究所 |
| 通讯作者 | Hu, ZH |
| 作者单位 | 1.Chinese Acad Sci, Wuhan Inst Virol, Joint Lab Invertebrate Virol, Wuhan 430071, Peoples R China 2.Chinese Acad Sci, Wuhan Inst Virol, Key Lab Mol Virol, Wuhan 430071, Peoples R China 3.Huazhong Agr Univ, Minist Agr, Key Lab Agr Microbiol, Wuhan 430072, Peoples R China |
| 推荐引用方式 GB/T 7714 | Hou, SW,Chen, XW,Wang, HZ,et al. An efficient method of constructing homologous recom binant baculovirus with pcr-amplified fragments[J]. Science in china series c-life sciences,2003,46(4):431-437. |
| APA | Hou, SW,Chen, XW,Wang, HZ,&Hu, ZH.(2003).An efficient method of constructing homologous recom binant baculovirus with pcr-amplified fragments.Science in china series c-life sciences,46(4),431-437. |
| MLA | Hou, SW,et al."An efficient method of constructing homologous recom binant baculovirus with pcr-amplified fragments".Science in china series c-life sciences 46.4(2003):431-437. |
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来源：武汉病毒研究所
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