Construction of a recombinant lentivirus expressing MCPH1 and establishment of its stably infected HeLa cell line

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Construction of a recombinant lentivirus expressing MCPH1 and establishment of its stably infected HeLa cell line

更新时间：2023-12-18

- Construction of a recombinant lentivirus expressing MCPH1 and establishment of its stably infected HeLa cell line
- Journal of Chongqing Medical University Vol. 38, Issue 3, Pages: 290-294(2014)
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- Published：2014，
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Construction of a recombinant lentivirus expressing MCPH1 and establishment of its stably infected HeLa cell line. [J]. Journal of Chongqing Medical University 38(3):290-294(2014)
DOI：

Construction of a recombinant lentivirus expressing MCPH1 and establishment of its stably infected HeLa cell line. [J]. Journal of Chongqing Medical University 38(3):290-294(2014) DOI：

摘要

目的：构建人MCPH1（microcephalin 1）基因重组慢病毒载体并建立稳定过表达MCPH1基因的宫颈癌HeLa细胞株。方法：将MCPH1基因重组到pLenO-DCE质粒中，构建重组质粒pLenO-DCE-MCPH1，经酶切和测序鉴定后，与慢病毒包装质粒共转染293T细胞，包装重组慢病毒，滴度测定后，感染HeLa细胞，用有限稀释法筛选稳定的细胞克隆，real-time PCR和Western blot分别检测细胞中MCPH1 mRNA和蛋白水平。结果：经酶切和测序鉴定，成功构建MCPH1重组慢病毒载体，并成功包装慢病毒，滴度为1.24×109 TU/ml；慢病毒感染HeLa细胞后经有限稀释法成功筛选到MCPH1稳定表达细胞株pLenO-DCE-MCPH1-HeLa；real-time PCR证实pLenO-DCE-MCPH1-HeLa细胞株较正常组HeLa（P=0.000）和pLenO-DCE-HeLa组（P=0.000）的MCPH1 mRNA水平显著地增高；Western blot进一步证实pLenO-DCE-MCPH1-HeLa细胞株过表达MCPH1。结论：成功构建MCPH1重组慢病毒，并建立了稳定表达MCPH1基因的细胞株pLenO-DCE-MCPH1-HeLa，为进一步研究MCPH1在宫颈癌中的作用奠定了基础。

Abstract

Molecular Medicine and Cancer Research Center；Teaching and Research Section of Biochemistry and Molecular Biology，College of Basic Medicine，Chongqing Medical University

关键词

慢病毒MCPH1基因宫颈肿瘤单克隆细胞

Keywords

lentivirusmicrocephalin 1 genecervical cancersingle clone cells

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