首页 | 本学科首页   官方微博 | 高级检索  
     

肝癌抗原肽(EPVTKAEML)与人HSP70融合基因的构建及原核表达
引用本文:曲萍,马加海,黄勇,刘利兵,铁茹,刘芳娥,黄小军. 肝癌抗原肽(EPVTKAEML)与人HSP70融合基因的构建及原核表达[J]. 细胞与分子免疫学杂志, 2008, 24(2): 113-114,118
作者姓名:曲萍  马加海  黄勇  刘利兵  铁茹  刘芳娥  黄小军
作者单位:1. 第四军医大学教学实验中心,陕西,西安,710032
2. 烟台毓璜顶医院麻醉科,山东,烟台,264000
3. 第四军医大学生物化学与分子生物学教研室,陕西,西安,710032
摘    要:目的:构建及原核表达肝癌抗原肽(EPVTKAEML)与人热休克蛋白70(HSP70)的融合基因.方法:采用加端PCR方法,将EPVTKAEML的基因序列融合到人HSP70基因的3′端;将融合基因克隆入原核表达载体pET-28a( )中,构建重组质粒pET-28a( )/EPVTKAEML-HSP70.经限制性内切酶BamH I、Xho I双酶切鉴定及序列测定后,转化E.coli BL21(DE3),经IPTG诱导表达融合蛋白,SDS-PAGE检测表达结果.结果:应用加端PCR方法扩增出约2.0 kb的目的片段,序列测定结果证实,EPVTKAEML的基因序列成功地融合到人HSP70基因的3′端.经BamH I、Xho I酶切鉴定证实,融合基因成功地克隆到原核表达载体pET-28a( )上;转入重组质粒的E.coli BL21(DE3)经IPTG诱导后,SDS-PAGE分析发现在相对分子质量(Mr)约72 000处有表达量明显增多的蛋白条带.结论:成功地构建并表达了肝癌抗原肽(EPVTKAEML)与人HSP70的融合基因.

关 键 词:肝肿瘤/免疫学  抗原  肿瘤/遗传学  表位  热休克蛋白类70/遗传学  基因表达  肝癌  抗原肽  融合基因  原核表达载体  heat shock protein  human  peptide  fusion gene  expression  白条  表达量  相对分子质量  发现  分析  基因克隆  测定结果  序列测定  应用  检测  融合蛋白
文章编号:1007-8738(2008)02-0113-03
收稿时间:2007-04-29
修稿时间:2007-06-01

Construction and expression of a fusion gene of human heptoma peptide with human heat shock protein 70
QU Ping,MA Jia-hai,HUANG Yong,LIU Li-bing,TIE Ru,LIU Fang-e,HUANG Xiao-jun. Construction and expression of a fusion gene of human heptoma peptide with human heat shock protein 70[J]. Chinese journal of cellular and molecular immunology, 2008, 24(2): 113-114,118
Authors:QU Ping  MA Jia-hai  HUANG Yong  LIU Li-bing  TIE Ru  LIU Fang-e  HUANG Xiao-jun
Affiliation:Center of Basic Medicine and Teaching Experiment, Fourth Military Medical University, Xirquote an 710032, China.
Abstract:AIM:To construct and express a fusion gene of human heptoma peptide (EPVTKAEML) with human heat shock protein 70 (HSP70). METHODS: A cDNA fragment encoding EPVTKAEML was added to 3 terminus of human HSP70 gene by PCR amplification. The PCR products of fusion gene were cloned into pET-28a(+)vector. The recombinant plasmid pET-28a(+)/EPVTKAEML-HSP70 was identified by enzyme digestion analysis and sequencing, and then it was transformed into E.coli BL21(DE3) through IPTG induction to express the target protein bearing His tag. RESULTS: A fragment of about 2.0 kb was amplified by PCR. Sequence analysis revealed that the sequence of EPVTKAEML was connected successfully to 3 terminus of human HSP70. Enzyme digestion analysis showed the fusion gene was cloned into pET-28a(+). SDS-PAGE showed that a relative molecular mass 72 000 fusion protein was expressed. CONCLUSION: The fusion gene of EPVTKAEML-HSP70 has been successfully constructed and expressed in E.coli BL21(DE3).
Keywords:hepatoma/immunology   antigen   neoplasm/ genetics   epitope   heat-shock protein 70/genetics   gene expression
本文献已被 维普 万方数据 等数据库收录!
设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号