Construction and expression of the eukaryotic expression plasmid of Human pEGFP/H2B

XU Yuan-jiu; et al

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Construction and expression of the eukaryotic expression plasmid of Human pEGFP/H2B

更新时间：2024-01-17

- Construction and expression of the eukaryotic expression plasmid of Human pEGFP/H2B
- Journal of Chongqing Medical University Vol. 34, Issue 8, Pages: 1060-1062(2009)
- 作者机构：
  
  1. 四川省广安市人民医院检验科
  2. 重庆市合川区中医院检验科
- 作者简介：
- 基金信息：
- DOI：
  CLC： R73-3
- Published：2009，
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[1]徐远久,熊异平,吴建敏,陈仕银,夏洪安,郑爱华,徐海波.人类pEGFP/H2B真核表达质粒的构建及表达[J].重庆医科大学学报,2009,34(08):1060-1062.

XU Yuan-jiu, et al. Construction and expression of the eukaryotic expression plasmid of Human pEGFP/H2B. [J]. Journal of Chongqing Medical University 34(8):1060-1062(2009)
[1]徐远久,熊异平,吴建敏,陈仕银,夏洪安,郑爱华,徐海波.人类pEGFP/H2B真核表达质粒的构建及表达[J].重庆医科大学学报,2009,34(08):1060-1062. DOI：

XU Yuan-jiu, et al. Construction and expression of the eukaryotic expression plasmid of Human pEGFP/H2B. [J]. Journal of Chongqing Medical University 34(8):1060-1062(2009) DOI：

摘要

目的:构建人H2B基因与GFP融合的绿色荧光蛋白真核表达质粒

并在HEK293细胞中表达。方法:通过RT-PCR从HEK293细胞总RNA中扩增获得H2B基因的部分cDNA序列

与载体pEGFP-C1连接构建H2B基因与GFP融合表达的真核表达质粒pEGFP/H2B

经酶切和测序鉴定;应用脂质体转染技术将重组质粒转染HEK293细胞观察融合蛋白的表达。结果:成功构建H2B基因与GFP融合的绿色荧光蛋白真核表达载体

与对照组中转染空载体pEGFP-C1的HEK293细胞中的弥漫荧光相比

转染重组质粒的细胞中可观察到绿色荧光蛋白集中表达在细胞核内

分裂期细胞中可见与染色体形态一致的绿色荧光。结论:该载体的成功构建为研究染色体的分离机制建立了有效的观察体系。

Abstract

Objective:To construct the eukaryotic expression plasmid of EGFP/H2B and express it in the Human embryo kidney cells(HEK293).Methods:Part H2B gene was obtained by RT-PCR from cultured HEK293 cell’s whole RNA.The sequence encoding H2B was cloned into pEGFP-C1 to construct the eukaryotic expression vector.After confirmed by enzymatic digestion and sequencing

the recombination plasmid was transfected to HEK293 cells by lipofectamine technology for observation of the fusion protein’s expression.Results:The recombination plasmid of pEGFP/H2B was constructed successfully.Compared with the dispersing green fluorescence among the whole cells in the control cells transfected with the mock plasmid(pEGFP-C1)

the green fluorescence was concentrated in the nuclei of HEK293 cells transfected with pEGFP/H2B and distributed with the chromosomes in the mitotic cells.Conclusions:The successful construction of pEGFP/H2B recombination plasmids establishes an available survey system to study the chromosome segregation mechanism.

关键词

H2B基因绿色荧光蛋白HEK293细胞染色体分离

Keywords

H2B genegreen fluorescence proteinHEK293 cellChromosome segregation

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