鸭IFN-α成熟肽基因的原核表达、复性及其抗病毒活性研究

畜牧兽医学报 ›› 2008, Vol. 39 ›› Issue (12): 1753-1758.doi:

鸭IFN-α成熟肽基因的原核表达、复性及其抗病毒活性研究

龚永强¹，程安春^1,2*，汪铭书^1,2*，杨梅¹，张瑶²，陈斌^1,3 ，钟小容¹

1.四川农业大学动物医学院禽病防治研究中心，雅安 625014；2.动物疫病与人类健康四川省重点实验室，雅安 625014； 3 四川省兽医防疫总站，成都 610041

收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2008-12-24 发布日期:2008-12-24
通讯作者: 程安春

Preliminary Studies on Prokaryotic Expression,Refolding of Duck IFN-α Mature Peptide and Its Activity against Virus

GONG Yong-qiang¹,CHENG An-chun^1,2*, WANG Ming-shu^1,2*,
YANG Mei¹, ZHANG Yao²,CHEN Bin^1,3,ZHONG Xiao-rong¹

1.Avain Disease Research Center of Sichuan Agricultural University, Ya’an 625014,China; 2.Key Laboratory of Animal Disease and Human Health of Sichuan Province, Ya’an 625014,China; 3.Sichuan Provincial Animal Epidemic Prevention & Supervision General Office，Chengdu 610041,China

Received:1900-01-01 Revised:1900-01-01 Online:2008-12-24 Published:2008-12-24

摘要/Abstract

摘要： 为获得鸭α-干扰素（DuIFN-α）并研究其生物学功能，将DuIFN-α成熟蛋白基因与原核表达载体pET32a+连接后转化到BL21（DE3）获得工程菌BL2l(DE3)/pET32a⁺-DuIFN-α，对其表达产物的纯化和复性关键技术及其抗水疱性口炎病毒（VSV）、鸭瘟强毒（DPV）活性进行研究。结果表明：BL2l(DE3)/pET32a⁺-DuIFN-α 经0.4 mmol/L IPTG诱导获得高效表达，表达的重组蛋白相对分子质量约37 ku，以包涵体形式存在；重组蛋白经镍金属螯合介质（Ni-MIAC）进行纯化和复性后获得理想效果，其抗VSV活性的比活力达到12 800 U/mg；利用定量PCR检测到15 U/mL的重组DuIFN-α对鸭瘟强毒表现出抑制作用，为重组DuIFN-α的临床应用提供试验数据。

Abstract: In order to obtain DuIFN-α and investigate its biological function, duck interferon-α(DuIFN-α) mature protein gene was cloned in to pMD18-T vector and inserted into the expressing plasmid/ pET32a⁺. Then,this recombinant plasmid was transformed into engineering bacteria/ BL2l(DE3) and a 37 kD protein was expressed in the form of inclusion bodies after 5 hours induction by 0.4 mmol/L IPTG. The expressed product which was purified and refolded by metal immobilization affinity chromatography(MIAC) have an ideal activity against virus. Its activity of anti-VSV in DEF was up to 12 800 U/mg and DuIFN-α at 15 U/mL can reduce duck plague virus replication by 80% as measured by FQ-PCR.

龚永强;程安春;汪铭书;杨梅;张瑶;陈斌;;钟小容. 鸭IFN-α成熟肽基因的原核表达、复性及其抗病毒活性研究[J]. 畜牧兽医学报, 2008, 39(12): 1753-1758.

GONG Yong-qiang;CHENG An-chun;WANG Ming-shu;YANG Mei;ZHANG Yao;CHEN Bin;ZHONG Xiao-rong. Preliminary Studies on Prokaryotic Expression,Refolding of Duck IFN-α Mature Peptide and Its Activity against Virus[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2008, 39(12): 1753-1758.

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