摘要
目的:合成抗血管生成肽βpep-25片断,构建并鉴定含βpep-25肽的重组原核表达质粒。方法:人工设计合成βpep-25肽序列片断,经测序正确后,利用基因克隆技术将其克隆到pGEM-Teasy载体中,构建重组质粒pGEM-T-βpep-25;用NaeI和BamHI双酶切pGEM-T-βpep-25后,将T-βpep-25肽片断克隆到经NaeI和BamHI双酶切的重组载体pBV220-NT4中,构建重组原核表达质粒pBV220-NT4-βpep-25,并对其分别用BamHI酶切,EcoRI和BamHI双酶切分析。结果:经酶切分析和DNA测序人工合成的肽βpep-25序列(113bp)与文献报道结果一致;分别用BamHI酶切,EcoRI和BamHI双酶切重组原核表达质粒pBV220-NT4-βpep-25可以得到4030bp、364bp和3666bp的片段,结果表明目的肽片段已经成功地克隆到了重组表达载体中,说明重组原核表达质粒构建成功。结论:抗血管生成肽βpep-25重组原核表达质粒的构建成功,为进一步在细胞内外及动物模型研究其作用机制和抗肿瘤作用奠定了基础。
AIM： To synthesize antiangiogenic peptide fragment of βpep-25, to construct and identify the recombinant prokaryotic expression plasmid containing βpep-25 peptide. METHODS： The fragment encoding βpep-25 peptide was designed and synthesized artificially and was cloned into vector pGEM-T easy after being identified by sequencing. After being digested by Nae I and BamH I, T-βpep-25 peptide fragment was cloned into recombinant vector pBV220NT4, which was digested by Nae I and BamH I. The constructed recombinant prokaryotic expression plasmid pBV220- NT4-βpep-25 was digested using BamH I, EcoR I and BamH I, respectively. RESULTS： The sequcence of βpep-25 synthesized artificially and analyzed by digestion was consistent with the published results. Fragments of 4 030 bp, 364 bp and 3 666 bp were obtained after digestion of recombinant prokaryotic expression plasmid pBV220-NT4-βpep-25 using BamH I, EcoR I and BamH I, respectively, which demonstrated the successful cloning and construction of recombinant prokaryotic expression plasmid. CONCLUSION： The successful construction recombinant prokaryotic expression plasmid expressing βpep-25 provides a foundation for further research on its mechanism of anti-tumor activity in vivo and in vitro.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2006年第2期154-156,160,共4页
Chinese Journal of Cellular and Molecular Immunology
基金
陕西省科学技术研究发展计划项目(No.2004K11-G3)
