全部 标题 作者
关键词 摘要

OALib Journal期刊
ISSN: 2333-9721
费用:99美元

查看量下载量

相关文章

更多...

猪γ干扰素的基因克隆、改造、表达及活性测定

DOI: 10.7685/j.issn.1000-2030.2003.02.017, PP. 71-75

Keywords: 猪γ干扰素,基因改造,原核表达,分离纯化

Full-Text   Cite this paper   Add to My Lib

Abstract:

提取经ConA诱导培养的猪外周血白细胞总RNA,RT-PCR扩增出猪IFN-γ基因并克隆到pGEM-T-easy载体,测序结果表明扩增片段是含有信号肽的猪IFN-γ的完整基因。设计1对引物亚克隆猪IFN-γ成熟蛋白基因并对其5′端前2个稀有密码子进行大肠杆菌偏嗜性改造。经测序鉴定后插入原核表达载体pRLC,并实现在大肠杆菌中的高效表达。表达产物以包涵体形式存在,经7mol?L-1盐酸胍的变性液溶解及05mol?L-1盐酸胍复性液复性处理,表达产物进行脱盐、凝胶层析纯化。细胞病变抑制法测定结果表明,重组猪IFN-γ具有较高的干扰素活性,约为45×106U?mg-1。

References

[1]  WilliamsJ G,MaierRV.Interferon-gamma: akeyimmunoregulatorylymphokine[J].Journal of Surgical Research.1993,54(1):79-93
[2]  张德震,吴淑华,侯云德.重组人γ干扰素的纯化[J].病毒学报.1989(1):3-8
[3]  张耀东,张丽华,耿信笃.人干扰素-γ基因工程大肠杆菌的培养及其产物的纯化与复性[J].生物工程学报.1999,15(2):240-243
[4]  Vandenbroeck K,Dijkmans R,Aerschot A V.Engineering by PCR - based exon amplification of the genomic porcine interferon-gam-ma DNA for expression in Escherichia coli[J].Biochemical and Biophysical Research Communications.1991,180(3):1408-1415
[5]  Bautista E M,Molitor T W.IFN -γ inhibits porcine reproductive and respiratory syndrome virus replication in macrophages[J].Archives of Virology.1999,144:1191-1200
[6]  Suradhat S,Intrakamhaeng M,Damrongwatanapokin S.The correlation of virus-specific interferon gamma production and protection against classical swine fever virus infection[J].Veterinary Immunology and Immunopathology.2001,83(3/4):177-189
[7]  Vandenbroeck K,Nauwynck H,Vancler pooten A.Recombinant porcine IFN-gamma potentiates the secondary IgG and IgA responses to an inactivated suid herpesvirus-1 vaccine and reduces post challenge weight loss and fever in pigs[J].Journal of Interferon & Cytokine Research.1998,18(9):739-744
[8]  HeathAW,Playfair H L.Conjugation of interferon-gamma to antigen enchances its adjuvanticity[J].IMMUNOLOGY.1990,71(3):454-456
[9]  Bao S,Beagley K W,France M P.Interferon plays a critical role in intestinal immunity against Salmonella typhimurium infection[J].IMMUNOLOGY.2000,99:464-472
[10]  陈德胜,潘杰彦,贾立群.传染性支气管炎病毒地方分离毒株S1基因的RT-PCR方法研究[J].南京农业大学学报.2000,23(3):85-88
[11]  李岩,邵念鹏,彭玉麟.γ干扰素的研究与临床应用(综述)[J].河北省科学院学报.2001,18(3):173-178
[12]  FARRAR M A,Schreiber R D.The molecular cell biology of interferon -γ and its receptor[J].Annual Review of Immunology.1993,11:571-61

Full-Text

Contact Us

service@oalib.com

QQ:3279437679

WhatsApp +8615387084133