鲤鱼中5个Sox基因保守区的克隆和比较

Cloning and comparison of conservative sequences of five Sox Genes in Cyprinus carpio

SRY/Sry基因已被公认为是哺乳动物的睾丸决定因子(TestisDeterminingFactor,TDF)基因,它的正确的时空表达是雄性生殖腺形成的关键,即导致哺乳动物胚胎性别决定的开关基因.作为一个大基因家族的首位成员,它的发现诱发了Sox基因家族的研究热潮.Sox基因家族是在动物中发现的一类新的编码转录因子的基因家族,其产物具有一个HMG基序保守区,参与诸如性别决定、骨组织的发育、血细胞生成过程、神经系统的发育、晶状体的发育等多种早期胚胎发育过程.鱼类是脊椎动物中进化地位较低的一类生物,除了个别种类出现了与性别相关的染色体外,绝大多数都无异形性染色体,说明了鱼类正处于性别染色体进化的重要时刻.研究鱼类中的Sox基因对于研究SRY的发生、性别染色体的进化以及性别的决定机制有着重要的意义.本实验利用兼并引物PCR的方法,参照Sox基因的HMG-box区氨基酸序列设计简并引物,对鲤鱼(Cyrinuscarpio)的基因组进行扩增,获得5个新的基因片段.经过在Genbank中进行同源性比较和分析,证明它们是鲤鱼的Sox基因并分别命名为CcSox3、CcSox4、CcSox11、CcSox14、CcSox21.与鲤鱼中的这些Sox基因具有最高同源性的基因分别是OlSox3,同源性为94.03%;CvSox4基因,同源性为88.06%;DrSox11基因,同源性为97.01%;MmSox14和HsSox14基?

In mammals, male sex determination is controlled by the presence of the Y chromosome, which carries a gene named SRY encoding the testis-determining factor (TDF). The product of this gene induces the testis-determining pathway, resulting ultimately in a male phenotype. SRY was found to contain a conservative HMG-box. A novel gene family SOX (SRY-related box) family was identified based on the conservation region of SRY. The Sox family of transcription factors are found throughout the animal kingdom, are characterized by the presence of a HMG domain, and are involved in the regulation of such diverse developmental processes of early embryogenesis as sex determination, chondrogenesis, haemopoiesis, neural development and lens development. Fishes serve as a key link on the view of the vertebrate systematic evolution(1). Its sex determination is primitive and plastically compared with the other advanced vertebrates. Sex chromosome differentiation in fishes is in the embryonic stage. Heteromorphic sex chromosomes have been found only in several species among the fishes that have been karyotyped. Sex determination type varies in different species. Extensive studies on the Sox genes of fishes will be helpful in approaching the evolution mechanism of sex-related genes and sex chromosomes. In this paper, Degenerate primers were designed on the basis of published Sox gene sequences among different species, which is special to the HMG-box motif. With the genomic DNA of Cyprinus carpio as template, one bands with the length 220bp were observed. Results of sequencing and Genbank database-searching showed that five novel gene fragment were obtained. The genes with the highest identity of possible amino acid sequence to these five novel gene fragments respectively are OlSox3 with identity 94.03%,CvSox4 with identity 88.06%, DrSox11 with identity 97.01%, MmSox14 and HsSox14 with identity 85.07%, HsSox21 with identity 95.52%. So .they were named as CcSox3、CcSox4、 CcSox11、 CcSox14、 CcSox21 respectively.