Molecular isolation and characterization of an expressed gene from the human Y chromosome
Molecular isolation and characterization of an expressed gene from the human Y chromosome
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DOI：
10.1093/hmg/1.9.717
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发表时间：
1992-01-01
影响因子：
3.5
通讯作者：
Lau, Yun-Fai Chris
中科院分区：
文献类型：
--
作者：
Zhang, Julie S.;Yang-Feng, Teresa L.;Lau, Yun-Fai Chris
利用定位克隆的方法，我们已经从流式分选的Y染色体粘粒文库中分离出了一个表达基因。该基因的分离是基于Y-231粘粒的鉴定，该粘粒在其人插入物中含有富含CpG的序列（HTF岛）。Y-231粘粒能检测人睾丸poly（A）+ RNA中1.3kb的转录本。从人睾丸cDNA文库中分离出几个cDNA克隆。此外，DNA介导的基因转移和限制性内切酶图谱实验表明，两个功能性转录单位内的Y-231粘粒。DNA序列分析表明，最大的cDNA克隆含有1075 bp的独特序列，并在相应mRNA的3'端含有一个poly（A）轨道。该基因的开放阅读框为762 bp，编码253个氨基酸，分子量为28.9 kD。Y-231结构基因包含约2.7 kb的基因组序列，并含有被5个内含子中断的6个外显子。Y-231基因在DNA水平上与先前描述的Y特异性基因睾丸特异性蛋白Y编码（TSPY）基因具有非常高（97%）的同一性，这表明这两个基因如果不相同，也可能相关。然而，TSPY基因被假定为无内含子的。进一步的PCR和RT-PCR分析这两个基因及其转录本提供了证据支持的假设，这两个基因是相同的基因，是一个Y-特异性的重复基因家族的成员含有内含子序列。Y-231（TSPY）基因在雄性基因组中是保守的，并在黑猩猩的睾丸中表达，这表明它可能在人类和类人猿的该器官的生理学中发挥重要作用。
Using a positional cloning approach, we have isolated an expressed gene from a flow-sorted Y chromosome cosmid library. The isolation of this gene was based on the identification of the Y-231 cosmid that contains CpG rich sequences (HTF islands) in its human insert. The Y-231 cosmid was capable of detecting a 1.3 kb transcript in poly (A)+ RNA samples from human testis. Several cDNA clones were isolated from a human testis cDNA library constructed in lambda-gt10. In addition, DNA-mediated gene transfer and restriction enzyme mapping experiments demonstrated that two functional transcriptional units are present within the Y-231 cosmid. DNA sequencing analysis showed that the largest cDNA clone contains 1075 bp of unique sequence and a poly (A) track at the 3' end of the corresponding mRNA. An open reading frame of 762 bp that encodes a predicted protein of 253 amino acids with a calculated molecular weight of 28.9 kD was identified. The Y-231 structural gene encompasses approximately 2.7 kb of genomic sequence and contains six exons that are interrupted by five introns. The Y-231 gene shares very high (97%) identity at the DNA level to a previously described Y-specific gene, testis specific protein Y-encoded (TSPY) gene, suggesting the possibility that these two genes are related, if not identical. However, the TSPY gene has been postulated to be intronless. Further PCR and RT-PCR analyses of these two genes and their transcripts have provided evidence supporting the hypothesis that they are the same gene and are members of a Y-specific repeated gene family containing intronic sequences. The Y-231 (TSPY) gene is conserved in the male genome and expressed in the testis of the chimpanzee, suggesting that it may play an important role in the physiology of this organ in man and the great ape.