摘要:作为男性生殖细胞，人类精子承担着将男性的遗传物质传递给下一代的重要任务。传统看法认为成熟精子是一个终末细胞，已经关闭基因的转录与翻译，但最近日益增多的研究资料，特别是芯片技术的广泛应用，发现成熟精子中存在有大量而复杂的mRNA 群体，然而其确切的种类和丰度尚不明确[1]。基因表达系列分析（SAGE）是一种不同于基因芯片的、具有数字化特征的基因表达分析方法，能从全基因组表达水平建立基因表达谱，具有易于克隆新基因的优势。本实验室已建立精子mRNA的SAGE库, 从该库中发现54个高丰度、但未能与数据库匹配的标签。本实验以这54个未匹配的SAGE标签作为研究的出发点，采用RT-PCR、Semi-nested PCR、5’-RACE等方法克隆出一条精子cDNA的3’-端片段。再结合生物信息学知识，获得了该基因的完整序列、开放阅读框及其染色体上的定位，检测其在一些器官组织中的表达情况发现该基因只在睾丸和精子中特异性表达，这说明该基因很可能与精子的发生或者生殖过程有关。对精子中未知基因的研究能更好的解读精子中RNA群体，精子RNA可能对诊断男性不育也具有非常大的应用潜力，因此对精子mRNA进行深入研究的意义重大。
关键词：人类成熟精子，mRNA，基因表达系列分析（SAGE），标签
Abstract:As male germ cells, the human spermatozoa take the important tasks of passing male germ plasm to the next generation. In traditional view, mature spermatozoa are highly differentiated terminal cells with genes' transcription and translation shut down, however, with the increasing number of recent research data, especially the wide application of chip technology, it is revealed that there exist a lot and yet complex popμLation of mRNA in mature spermatozoa with their exact type and abundance unclear. Serial Analysis of Gene Expression(SAGE) , different to gene chip, is a gene expression analysis method with digital characteristic, it can establish gene expression profile in the level of genome expression, and it has an advantage of cloning new genes easily. Our lab has established SAGE library of human mature spermatozoa mRNA, and in this library we found 54 SAGE high-abundanced tags that can't match in database. In this experiment, starting from those 54 unmatched SAGE tags, we used methods of RT-PCR, Semi-nested PCR and5’-RACE etc. to clone a 3'-fragment of sperm cDNA.Then using knowledge of bioinformatics, we obtained the complete sequences, open reading fragment, and position on chromosomes of this gene. We found that this gene was specially expressed in testicle and sperm through checking the expression in tissues and organs, it shows that this gene probably have relation to spermatogenesis and production process.It can read RNA group in sperm better to research on unknown genes in sperm,and perm mRNA may have a great application potential in the diagnosis of male contraception, so it makes great sense to do deeper research on sperm mRNA
Key words: Human mature spermatozoa, mRNA, Serial Analysis of Gene Expression(SAGE), tag
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