应用基因芯片技术研究成年男性精子的基因表达

Research of the Spermatozoal Gene Expression with Gene Microarrays

目的:初步研究健康成人精子中基因的表达情况。方法:收集成年男性活力正常的精子,提取总RNA,以健康成人淋巴细胞总RNA作对照。两组RNA逆转录为cDNA之后,cDNA经酶切、加接头后进行RD扩增,在扩增同时分别掺入Cy3(精子)和Cy5(淋巴细胞)荧光染料标记,标记样品与自制的精子基因表达谱芯片杂交。结果:检测发现在560个基因片段中,有72个基因表达上调,321个基因表达下调,另外167个基因表达无差异。其中与基因复制、转录、翻译及调控等相关的基因表达基本属于无差异表达类型或低表达类型,而与精子发生相关的基因、精子本身的特异性抗原等基因显著高表达,精子中糖酵解相关基因表达上调,而与氧化磷酸化相关的基因则表达下调。结论:在健康成人精子中有丰富的基因表达,而且基因的表达与精子自身的功能和特点相一致。

Objective： To perform the detection of spermatozoal gene expression in order to accelerate the study of spermatozoal molecular biology. Methods： To collect the healthy adults＂ sperm and lymphocytes respectively, and then to extract the total RNAs from them by RNeasy mini kit（QIAGEN） or Trizol reagent. Corresponding cDNAs were produced, digested, ligated, finally labeled with Cy3 （sperm） and Cy5 （lymphocyte） in the course of RD amplifying reactions. Hybridization with self-made micrarrays contained 560 probes was carried out after the labeled cDNAs pured by PCR Product Purification Kit. Results： Among the 560 probes, 72 genes were up-regulated, 321 genes were down-regulated, the others had no different expression. Furthermore, genes associated with replication, transcription, translation and regulative functions were non-different expression or down-regulated, and those belonged to the spermatogenesis associated, sperm associated antigen were up-regulated, but those involved in the glycolysis were up-regulated, in the oxidative phosphorylation were down-regulated. Conclusion ： It had successfully confirmed that there were a plenty of genes expressed in sperm, furthermore the genes expressed were accorded to spermatozoal functions and characteristics.