贺超 洪广 吴静娴 冯正阳 王文基 陈华谱 朱春华 李广丽

摘要[目的]克隆性别调控基因Sox9的cDNA序列，为后续开展Sox9分子进化及性别调控功能的研究提供参考。[方法]从泰国斗鱼中克隆鉴定出Sox9基因的cDNA序列全长，并展开序列分析及在雌雄个体中的组织分布研究。[结果]利用Smart-RACE的分子克隆方法，从泰国斗鱼精巢中克隆得到Sox9基因的cDNA序列，全长为2 201 bp，其中开放读码框1 449 bp；氨基酸比对分析显示Sox9氨基酸序列中的HMG序列的保守性相当高；通过系统进化树分析发现泰国斗鱼Sox9与半滑舌鳎的亲缘关系最近；利用RT-PCR的方法检验了泰国斗鱼Sox9基因在雌雄个体中的组织分布，结果发现泰国斗鱼Sox9基因在雌性的肌肉中表达量最高，其次在脑和肠中，而在卵巢未检测到表达信号；在雄性中，Sox9基因在脑和精巢中的表达量最高。[结论]泰国斗鱼Sox9具有保守的结构特征，并且在组织分布中显示出表达特异性及性别差异性。

关键词泰国斗鱼；Sox9；序列分析；组织分布

中图分类号S917.4文献标识码

A文章编号0517-6611（2017）08-0151-05

Study on Cloning and Tissue Expression of Sox9 in Betta splendens

HE Chao，HONG Guang，WU Jingxian，CHEN Huapu* et al（Key Laboratory of Marine Ecology and Aquaculture Environment of Zhanjiang，Guangdong Ocean University，Zhanjiang，Guangdong 524088）

Abstract[Objective] To clone the fulllength cDNA sequence of sexual regulation gene Sox9 ，to provide a basis for the molecular evolution and functional studies of Sox9 in Betta splendens.[Method]The Sox9 cDNA sequence in B.splendens was cloned.Sequence analysis and research of organizations distribution in the male and female individual was carried out.[Result]The Sox9 cDNA sequence in the testis of B.Splendens was cloned by using the rapidamplification of cDNA ends （RACE），it contained 2 201 bp nucleotides with the open reading frame （ORF） of 1 449 bp；the amino acid sequence alignment showed that the HMG （the high mobility group box） domain of Sox9 was rather conserved； based on the phylogenetic analysis，the B.splendens Sox9 protein was closer to Cynoglossus semilaevis；tissue distribution analyzed by RTPCR showed that the expressions of Sox9 were highly expressed in muscle，moderate in brain and gun，and undetectable in ovary in female； high expression level of Sox9 was found in testis and brain in male.[Conclusion]The Sox9 in B.Splendens shows conservative structural characteristics，tissue express specificity and gender differences.

Key wordsBetta splendens；Sox9；Sequence analysis；Tissue distribution

Sox（SRYrelated high mobility group box）家族目前已經被克隆鉴定出30多个成员。Sox家族成员均含有79个高度保守HMG盒（high mobility group box）的结构域，主要是与下游调控基因的DNA结合，成为Sox家族成员的最显著性结构特征[1]。其中Sox9被认为是Sox家族中与性别分化较相关的成员之一[2]，受到了广泛的关注与研究。在哺乳类、鸟类和爬行类的性分化时期，Sox9基因在雄性生殖嵴的表达量急剧上调[3-5]。在小鼠中，雌性性腺中Sox9的异常表达导致了睾丸的形成[6]。Sox9基因的缺失会造成雄性逆转成为雌性，从而表明Sox9基因在精巢发育中起了至关重要的作用，并被认为是雄性性别决定因子[7]。……