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学科主题: 基础医学
题名:
Generating DNA sequences encoding tandem peptide repeats suitable for expression and immunological application
作者: Hou, Hongwei1; Zhang, Zhiqian1; Zhao, Wei1; Hou, Jinyan2
关键词: Tandem repeats ; Peptide ; Ligation-PCR ; Peptide production ; Antibody
刊名: WORLD JOURNAL OF MICROBIOLOGY & BIOTECHNOLOGY
发表日期: 2012-05-01
DOI: 10.1007/s11274-012-1023-4
卷: 28, 期:5, 页:2175-2180
收录类别: SCI
文章类型: Article
WOS标题词: Science & Technology
类目[WOS]: Biotechnology & Applied Microbiology
研究领域[WOS]: Biotechnology & Applied Microbiology
关键词[WOS]: POLYMERASE CHAIN-REACTION ; DIRECT CLONING ; VACCINE DESIGN ; PCR PRODUCTS ; CONTAINING VECTOR ; CANCER
英文摘要:

Tandem repeats of single short peptide sequences are useful for many purposes. Here we describe a method called ligation-PCR to construct DNA sequences encoding numerous tandem peptide repeats that can stably produce such repeats in both prokaryotic and eukaryotic cells. The method employs double-strand target monomers consisting of a short peptide coding sequences. These sequences contain 3-bp cohesive overhangs to ensure correct repeat orientation and reading frame during ligation. The ligation products are PCR amplified and directly cloned into a new TA-cloning vector, pZeroT. Constructs containing tandem 10-amino-acid myc-tag peptide coding sequence repeats that ranged from approximately 0.45-1.2 kb, representing 15-40 copies of the corresponding peptide, were successfully obtained by this method. When one of the constructs was subcloned into prokaryotic vector pET-28 c (+) and eukaryotic vector rGHpcDNA3.0, and introduced into E. Coli and COS-7 cells, respectively, proteins containing tandem myc-tag peptide repeats were expressed with expected molecular weights. Purified proteins from E. Coli could successfully stimulate a peptide specific immune response. This method provides a means to manipulate peptides at the nucleic acid level, and can serve as the basis for biological peptide synthesis, epitope-specific antibody production, and epitope-based DNA vaccine construction.

语种: 英语
所属项目编号: 39880016 ; 30971494 ; 2011-2-24 ; 7972010 ; 5122012
项目资助者: National Natural Science Foundation of China ; Beijing Municipal Outstanding Talents Training Funds in Health Sciences ; Beijing Municipal Natural Science Foundation
WOS记录号: WOS:000303385900034
Citation statistics:
内容类型: 期刊论文
URI标识: http://ir.bjmu.edu.cn/handle/400002259/65084
Appears in Collections:基础医学院_细胞生物学系_期刊论文

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作者单位: 1.Peking Univ, Canc Hosp & Inst, Dept Cell Biol, Key Lab Carcinogenesis & Translat Res,Minist Educ, Beijing 100142, Peoples R China
2.Qingping Bio Co Ltd, Toronto, ON M3H 1X8, Canada

Recommended Citation:
Hou, Hongwei,Zhang, Zhiqian,Zhao, Wei,et al. Generating DNA sequences encoding tandem peptide repeats suitable for expression and immunological application[J]. WORLD JOURNAL OF MICROBIOLOGY & BIOTECHNOLOGY,2012,28(5):2175-2180.
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