Cloning,expression and purification of human canstatin and assay of its bioactivity

ZHANG Xiangrong; et al

doi:10.13406/j.cnki.cyxb.2005.04.004

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Cloning,expression and purification of human canstatin and assay of its bioactivity

更新时间：2024-01-17

- Cloning,expression and purification of human canstatin and assay of its bioactivity
- Journal of Chongqing Medical University Issue 4, Pages: 497-500(2005)
- 作者机构：
  
  1. 重庆医科大学基础医学院生物化学教研室
  2. 重庆医科大学基础医学院生物化学教研室,重庆,400016
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.2005.04.004
  CLC： Q78
- Published：2005，
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[1]张向荣,孙厚良,曾昭淳.人canstatin基因的克隆、表达、纯化及其生物活性测定[J].重庆医科大学学报,2005(04):497-500.

ZHANG Xiangrong, et al. Cloning,expression and purification of human canstatin and assay of its bioactivity. [J]. Journal of Chongqing Medical University (4):497-500(2005)
[1]张向荣,孙厚良,曾昭淳.人canstatin基因的克隆、表达、纯化及其生物活性测定[J].重庆医科大学学报,2005(04):497-500. DOI： 10.13406/j.cnki.cyxb.2005.04.004.

ZHANG Xiangrong, et al. Cloning,expression and purification of human canstatin and assay of its bioactivity. [J]. Journal of Chongqing Medical University (4):497-500(2005) DOI： 10.13406/j.cnki.cyxb.2005.04.004.

摘要

目的:克隆人Canstatin cDNA

在E.coli中融合表达和纯化

并测定表达产物抑制新生血管生成活性。方法:从中国人胎盘组织提取总RNA

RT-PCR扩增出Canstatin cDNA

克隆入pTYB1载体中并测序

构建原核表达载体pTYB1-hCAN

IPTG诱导表达

几丁质亲和纯化

鸡胚绒毛尿囊膜（CAM）血管生成实验进行活性测定。结果:RT-PCR产物约为700bp

测序结果与文献报道序列一致。构建了人Canstatin原核表达载体pTYB1-hCAN

在大肠杆菌中表达。纯化的融合蛋白在CAM实验中能明显抑制血管生成。结论:克隆、表达了具有生物学活性的人canstatin蛋白。

Abstract

Objective:To clone human canstatin cDNA

express it as fusion protein in E.coli

purify the fusion protein and determine its biological activity.Methods:Total RNA was extracted from Chinese placenta tissue.The canstatin cDNA was amplified by RT-PCR

then cloned into pTYB1 and sequenced;Prokaryotic expression vector pTYB1-hCAN was constructed;fusion protein was expressed by using IPTG induction and purified by chitin column.The biological activity of the purified fusion protein was identified by using the chick embryo chorioallantoic membrane assay(CAM).Results:RT-PCR product was about 700 bp

its sequence was the same as that of canstatin reported.Prokaryotic expression vector pTYB1-hCAN of human canstatin was constructed successfully

and the fusion canstatin protein was expressed in E.coli.The purity of the protein was 91.6%.The fusion protein could suppress the angiogenesis in CAM.Conclusion:The active human Canstatin is cloned and expressed as a bioactive form.

关键词

canstatin克隆基因表达蛋白质纯化抗血管生成

Keywords

Canstatinclonegene expressionProtein purificationanti-angiogenesis

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