梅花鹿IFN-β1的原核表达及活性鉴定
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- 英文论文题名：Study on prokaryotic expression of interferon-β1 mature peptide gene of Sika deer
- 论文作者：苏凤艳 ; 李哲 ; 刘存发 ; 宗颖 ; 曾范利 ; 王全凯
- 英文论文作者：Su Fengyan ; Li zhe ; Liu Chunfa ; Zong Ying ; Zeng Fanli ; Wang Quankai ; College of Chinese Medicinal Materials ; Ji Lin Agricultural University ; Wild Animal rescue and Breed center of Jilin Province ; Jinlin Sino-ROK Academy of Animal Sciences
- 年：2016
- 作者机构：吉林农业大学中药材学院;吉林省野生动物救护繁育中心;吉林省中韩动物科学研究院;
- 论文关键词：梅花鹿 ; IFN-β1基因 ; 原核表达 ; 活性鉴定
- 英文论文关键词：sika deer ; IFN-β1 gene ; prokaryotic expression ; activity identification
- 会议召开时间：2016-06-01
- 会议录名称：“十二五” 鹿科学研究进展
- 语种：中文
- 分类号：S858.25
- 学会代码：ZXMY
- 学会名称：中国畜牧业协会
- 页数：7
- 文件大小：721k
- 原文格式：O
摘要
为表达具有抗病毒活性的梅花鹿干扰素-β1(IFN-β1),本研究通过PCR扩增梅花鹿IFN-β1基因,将其克隆至原核表达载体pET-28a中,构建重组质粒pETIFNβ1,并将其转化至大肠杆菌Rosetta(DE3)感受态细胞中,经IPTG诱导后,得到高效表达。SDS-PAGE与western blot检测结果表明,重组梅花鹿IFN-β1重组蛋白的分子量大小约为24 ku,表达产物主要为不溶性的包涵体,表达量占菌体总蛋白的59.02%。将Ni柱纯化的pETIFNβ1诱导表达产物复性后,细胞病变抑制法检测表明表达产物具抗病毒活性。梅花鹿INF-β1基因的表达,为梅花鹿INF-β1的应用奠定了基础。
In order to express the sika deer interferon batal(IFN- β1) with antiviral activity,IFN- β1 gene was amplified by PCR.Then the gene was inserted into prokaryotic expression vector pET28 a to eontruet recombinant plasmid pETIFNβ1.pETIFNβ1 was transformed into E.coli Rosseta(DE3).The recombinant protein was expressed in E.coli Rosseta(DE3)by IPTG inducing.The expression product was identified by SDS- PAGE and Western-blot.The results showed that IFN- β1 recombinant protein of Sika deer was about 24 Ku and mainly for the insoluble inclusion body.The expression product was account for 59.02%of the total bacteria protein.The expression product of pETIFNβ1 was purified by the Ni column and renatured.Then,cytopathic inhibition method detected that expression product possess antiviral activity.This study set a basis for future application of sika deer IFN-β1.
In order to express the sika deer interferon batal(IFN- β1) with antiviral activity,IFN- β1 gene was amplified by PCR.Then the gene was inserted into prokaryotic expression vector pET28 a to eontruet recombinant plasmid pETIFNβ1.pETIFNβ1 was transformed into E.coli Rosseta(DE3).The recombinant protein was expressed in E.coli Rosseta(DE3)by IPTG inducing.The expression product was identified by SDS- PAGE and Western-blot.The results showed that IFN- β1 recombinant protein of Sika deer was about 24 Ku and mainly for the insoluble inclusion body.The expression product was account for 59.02%of the total bacteria protein.The expression product of pETIFNβ1 was purified by the Ni column and renatured.Then,cytopathic inhibition method detected that expression product possess antiviral activity.This study set a basis for future application of sika deer IFN-β1.
引文
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[2]Sin WX,Li P,Yeong JP,et al.Activation and regulation of interferon-βin immune responses[J].Immunol Res,2012,53(1-3):25-40.
[3]曲嘉琪,李洋,钟一鸣,等.β-干扰素及其研究进展[J].安徽农业科学,2013,41(9):3792-3793,3582.
[4]郑拓,吴长德.犬干扰素的研究进展[J].中国兽医杂志,2012,48(1):77-79.
[5]张译元,张廷红,常维山.水貂β-干扰素基因的克隆、测序与遗传进化分析[J].中国兽药杂志,2012,46(1):10-12.
[6]姜亦飞,李峰,张世栋,等.我国梅花鹿产业的发展现状及前景展望[J].山东农业科学,2012,44(9):109-111,114.
[7]吕见涛,张一斌,陈法荣,等.梅花鹿口蹄疫免疫操作技术[J].中国兽医杂志,2010,46(1):47-48.
[8]杜锐,尹茉莉,时坤,等.抗鹿源牛病毒性腹泻—粘膜病病毒特异性单克隆抗体的制备与鉴定[J].中国预防兽医学报,2010,32(8):641-643.
[9]Diebold S S,Montoya M,Unger H,et al.Viral infection swit-ches non-plasmacytoid dendritic cells into high interferon pro-ducers[J].Nature,2003,424(6946):324-328.
[10]张永.干扰素的分子生物学机制研究进展[J].海峡药学,2007,19(1):7-10.
[11]Low-Calle AM,Prada-Arismendy J,Castellanos JE.Study of interferon-βantiviral activity against Herpes simplex virus type 1 in neuron-enriched trigeminal ganglia cultures[J].Virus Res.2014,180:49-58.
[12]Koerner I,Koghs G,Kalinke U,et al.Protective role of interferon-βin host defense against influenza A virus[J].J Virol,2006,10:1-15.
[13]Hensley L E,Fritz E A,Jahrling P B,et al.Interferon-β1a and SARS Coronavirus Replication[J].Emerging Infectious Diseases,2004,10:317-319.
[14]CAI MH,ZHU F,SHEN PP.Expression and purification of chicken beta interferon and its antivirus immunological activity[J].Protein Expr Purif,2012,84;123-129.