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人硫氧还蛋白cDNA的克隆、表达及其多克隆抗体制备 被引量：1

Cloning and Expression of Human Thioredoxin-1 cDNA and Preparation of Its Polyclonal Antibody
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摘要 目的制备硫氧还蛋白1(thioredoxin-1,Trx-1)多克隆抗体。方法从乳腺癌细胞系MCF-7中用RT-PCR的方法得到了Trx-1全长基因,将它克隆到原核表达载体上进行大量的表达和纯化,纯化的蛋白对新西兰大白兔进行背部多点注射,40d后取其血清用梯度饱和硫酸铵沉淀的方法进行多克隆抗体的纯化。用ELISA和Western印迹实验测定抗体效果。结果成功获得了Trx-1全长cDNA,通过原核表达得到了大量Trx-1蛋白,并制备了高效价的多克隆抗体。结论此多克隆抗体对Trx-1蛋白具有良好的识别能力,可以应用于Trx-1的功能研究。 Objective To obtain the Trx-1 polyclonal antibody. Methods cDNA encoding the Trx-1 from MCF-7 cells was isolated by RT-PCR method. The Trx-1 protein expressed by pET32a vector and purified was injected into rabbits to prepare polyclonal antibody. Western blotting and ELISA were used to detect the effect of the polyclonal antibody. Results Trx-1 was successfully ob- tained and was successfully be expressed and purified. Its polyclonal antibody was also obtained. Conclusion Trx-1 protein was well reactive to its polyclonal antibody. The polyclonal antibody may be used for the study of the physiological function of the Trx-1 protein.
作者 陈孝平 唐文心 陈欢 付春华
机构地区 华中科技大学生命科学与技术学院生物物理研究所
出处 《医学分子生物学杂志》 CAS CSCD 2007年第3期221-224,共4页 Journal of Medical Molecular Biology
关键词 硫氧还蛋白 CDNA克隆 多克隆抗体 thioredoxin cDNA clone polyclonal antibody
分类号 Q7 [生物学—分子生物学]
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参考文献12

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同被引文献10

  • 1POWIS G,MONFFORT W R. Properties and biological activ ities of thioredoxins[J]. Annu Rev Biophys Biomol Struct, 2001,30:421-455.
  • 2ARNOLD N B, KETTERER K, KLEEFF J. Thioredoxin is downstream of smad7 in a pathway that promotes growth and suppresses cisplatin induced apoptosis in pancreatic cancer [J]. Cancer Res, 2004,15(64):3599-3606.
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  • 6YOUNG M G, THOMAS R Z, JENNIFER M J, et al. Selective protection of nuclear thioredoxin 1 and glutathione redox systems against oxidation during glucose and glutamine deficiency in human colonic epithelial cells[J]. Free Radic Biol Med, 2007,42(3) :363-370.
  • 7UENO M, MATSUTANI Y, NAKAMURA H, et al. Pos sible association of thioredoxin and p53 in breast cancer[J]. Immunol Lett, 2000,75(1) :15-20.
  • 8LI Y X, LIU W Y, XING G C, et al. Direct association of hepatopoietin with thioredoxin constitutes a redox signal transduction in activation of AP-1/NF-κB[J]. Cell Signal, 2005,17(8) :985-996.
  • 9RAVI D, MUNIYAPPA H, KUMUDA C D. Endogenous thioredoxin is required for redox cycling of anthracyclines and p53 dependent apoptosis in cancer cells[J]. J Biol Chem, 2005, 280(48) :40084-40096.
  • 10CHEN J, COUTO F M, MINN A H, et al. Exenatide inhibits b-cell apoptosis by decreasing thioredoxin interacting protein[J]. Biochem Biophys Rts Commun, 2006, 346(3):1067-1074.

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医学分子生物学杂志
2007年 第3期

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