染色体数目异常滋养层细胞中CENP-I基因的的异常表达

目的:探讨人胚胎滋养层细胞中染色体数目异常与着丝粒特异性蛋白质CENP-I表达水平的相关性。方法:T-A克隆制备CENP-I、GAPDH定量PCR标准品;用Ni2+-NTA亲和层析柱纯化CENP-I重组蛋白,免疫兔制备抗人CENP-I多克隆抗体;采用FQ-PCR和Western-blot检测23例染色体数目异常的滋养层细胞(实验组)和30例具有正常核型的滋养层细胞(对照组)中CENP-ImRNA和蛋白表达水平。结果:成功制备CENP-I、GAPDHFQ-PCR标准品,表达和纯化CENP-I重组蛋白,获得兔抗人CENP-I多克隆抗体;FQ-PCR和Western-blot结果显示CENP-I在实验组和对照组中均有表达,但实验组中CENP-I的表达水平明显降低(P<0.05)。结论:CENP-I表达的降低可能是导致人胚胎滋养层细胞中染色体数目异常的原因之一。

Abnormal Expression of cENP-Gene in Trophoblast Cells with Numerical Chromosomal Aberration

Objective:To observe the expression level of CENP-I gene in human trophoblast cells with numerical chromosomal aberration. Methods:Recombinant plasmids pMD18-T/CENP-I and pMD18-T/GAPDH were constructed and used as the standard plasmids in quantitative real-time PCR. Recombinant CENP-I protein were expressed in E. coli and purified using the Ni2+-NTA affinity, and subsequently used as the immunogen to prepare the rabbit anti-CENP-I antibody. Quantitative real-time PCR and Western-blot were used to evaluate the endog-enous expression level of CENP-I mRNA and CENP-I protein in human trophoblast cells. Results:pMD18-T/ CENP-I and pMD18-T/CENP-I standard plasmids of real-time PCR were constructed, and the rabbit anti-human CENP-I polyclone antibody was prepared. The expression of CENP-I gene were detected both in normal trophoblast cells and trophoblast cells with numerical chromosomal aberration;however, the expression levels of CENP-I mRNA and CENP-I protein were significantly decreased in trophoblast cells with numerical chromosomal aberra-tion compared to that in normal trophoblast cells. Conclusion:The down-regulated expression of CENP-I gene may be one of the reasons that result the abnormal number of chromosome in human trophoblast cells.