期刊文献+

TGF-β1和TGF-βⅡ型受体酵母表达载体构建 被引量:3

Construction and identification of yeast two-hybrid expression vector containing human transforming growth factor-beta 1 and different fragments of transforming growth factor-beta receptor type Ⅱ
原文传递
导出
摘要 目的克隆人转化生长因子β1(TGF-β1)和4个受体基因片段,构建酵母表达载体,为应用酵母双杂交筛选与TGF-β1相互作用的TβRⅡ区域提供技术支持。方法提取人血液中RNA,逆转录转为c DNA,PCR扩增TGF-β1和4个受体(TβRⅡ-A、TβRⅡ-B、TβRⅡ-C、TβRⅡ-D)基因片段,双酶切后,分别插入酵母双杂交表达载体p GBKT7和p GADT7;经过PCR扩增、双酶切和DNA测序鉴定后,构建质粒p GBKT7-TGF-β1和p GADT7-TβRⅡ-A、p GADT7-TβRⅡ-B、p GADT7-TβRⅡ-C和p GADT7-TβRⅡ-D。结果成功扩增TGF-β1基因(1 200 bp)和TβRⅡ-A(453bp)、TβRⅡ-B(366 bp)、TβRⅡ-C(276 bp)和TβRⅡ-D(165 bp)基因片段,经PCR、双酶切和DNA测序鉴定插入到酵母双杂交载体序列正确。结论成功构建p GBKT7-TGF-β1和p GADT7-TβRⅡ-A、p GADT7-TβRⅡ-B、p GADT7-TβRⅡC和p GADT7-TβRⅡ-D酵母双杂交表达载体。 Objective To clone human transforming growth factor-beta 1 (TGF-β1) and 4 genome fragments of its receptor and to construct the yeast expression vector to provide technical support for using yeast two-hybrid in identification of interaction domain of TGF-β type Ⅱ receptor (TβRⅡ) with TGF-β1.Methods The gene TGF-β1 and gene fragments of TβRⅡ from the RNA of human peripheral blood were amplified with regular PCR using specific primers,and then cloned into yeast two-hybrid expression vector pGBKT7 and pGADT7.The recombinant plasmids pGBKT7-TGF-β1 and pGADT7-TβRⅡ-A,pGADT7-TβRⅡ-B,pGADT7-TβRⅡ-C,pGADT7-TβRⅡ-D sequences were confirmed with PCR,restriction endonucleases digestion,and automated DNA sequencing.Results The yeast two-hybrid expression vector pGBKT7-TGF-β1 (1 200 bp) and pGADT7-TβRⅡ-A (453 bp),pGADT7-TβRⅡ-B (366 bp),pGADT7-TβRⅡ-C (276 bp),pGADT7-TβRⅡ-D (165 bp) were successfully constructed and the sequences of the plasmids’ DNA were correct.Conclusion The yeast two-hybrid expression vectors pGBKTT-TGFβ1 and pGADT7-TβRⅡA,B,C,and D were successfully constructed.
出处 《中国公共卫生》 CAS CSCD 北大核心 2017年第6期926-929,共4页 Chinese Journal of Public Health
基金 国家自然科学基金(81500471) 黑龙江省自然科学基金(H2015080) 黑龙江省卫生厅项目(2014-212 2016-355) 研究生创新科研基金(2014YJSCX-07)
关键词 人转化生长因子β1(TGF-β1) TGFβⅡ型受体 质粒构建 酵母双杂交 transforming growth factor-beta 1 transforming growth factor-beta receptor type Ⅱ plasmid construc-tion yeast two-hybrid
  • 相关文献

参考文献2

二级参考文献32

  • 1周广彪,胡盛平,刘静文,张可浩,官兵,吴名耀,沈忠英,杨捷生.改良手工显微切割法获取特定细胞提取高质量RNA[J].中国生物化学与分子生物学报,2006,22(3):252-257. 被引量:4
  • 2黄平,柯昌文,邹丽容,李晖,陈秋霞.广东地区人禽流感H_5N_1血凝素基因特征与进化[J].中国公共卫生,2007,23(4):390-392. 被引量:11
  • 3J.莎姆布鲁克(著),黄培堂(译).分子克隆实验指南[M].3版.北京:科学出版社,2002:1428-1480.
  • 4Sym D,Patel PN,EI-Chaar GM. Seasonal,avian,and novel H1N1 influenza: prevention and treatment modalities [ J ]. Ann Pharmacotlaer,2009,43 (12) :2001 - 2011.
  • 5Mitrasinovic PM. Advances in the structure-based design of the influenza A neuraminidase inhibitors[J]. Curr Drug Targets. 2010, 11 (3) :315 -326.
  • 6Li Z, Xiong F, Lin Q, et al. Low-temperature increases the yield of biologically active herring antifreeze protein in Pichia pastoris [J]. Protein Expr Purif,2001,21 (3) :438 -445.
  • 7Cregg JM ,Cereghino JL, Shi J, et al. Recombinant protein expression in Pichia pastoris[J]. Mol Biotechnol,2000,16 ( 1 ) :23 - 52.
  • 8Keitel WA,Atmax RL.Preparing for a possible pandemic: influenza A/H5N1 vaccine development [ J ]. Curr Opin Pharmanol, 2007,7(5) :484 -490.
  • 9Stiver HG. The threat and prospects for control of an influenza pandemic[ J]. Expert Rev Vaccines,2004,3 ( 1 ) :35 -42.
  • 10Tjoa ML,Jani J,Lewi L,et al.Circulating cell-free fetalmessenger RNA levels after fetoscopic interventions ofcomplicated pregnancies[J].Am J Obstet Gynecol,2006,195(1):230-235.

共引文献3

同被引文献10

引证文献3

相关作者

内容加载中请稍等...

相关机构

内容加载中请稍等...

相关主题

内容加载中请稍等...

浏览历史

内容加载中请稍等...
;
使用帮助 返回顶部