黄鳝WT1基因序列分析及在性腺发育过程中的表达

Sequence analysis and expression of the Wilms' tumor suppressor gene(WT1)during sex reversal in rice field eel,Monopterus albus

采用RT-PCR法从黄鳝(Monopterus albus)性腺mRNA中克隆获得了WT1基因,根据其氨基酸序列有无KTS区将黄鳝WT1基因分为2种类型:WT1(-KTS)为1 236bp,WT1(+KTS)为1 245bp。与NCBI/GenBank上其他物种进行氨基酸序列比对,发现它们与斜带石斑鱼的WT1a相似性最高,分别为93.2%和93.0%,并且它们均在羧基端具有相似性很高的锌指结构区域。系统进化树分析发现,黄鳝WT1基因与斜带石斑鱼WT1a聚类为一支。WT1基因在黄鳝性腺、肾、肠、脾和心脏组织中都有表达,且该基因在黄鳝雌性、间性和雄性性腺中的表达量在各时期都有先升高后降低的表达趋势。

Two major isoforms,with an insertion （＋KTS） or exclusion （-KTS） in amino acid se- quences, resulting from alternative splicing, were cloned from the gonad of rice field eel Monopterus albus by RT-PCR. The length of cDNA sequence of the two WT1 isoforms, WT1 （-KTS） and WT1 （＋ KTS）,were 1 236 bp and 1 245 bp, respectively. The results showed that the amino acid sequences of WT1 （-KTS） and WT1 （＋KTS） in M. albus shared 93.2% and 93.0% identity with WTla in Epi- nephelus coioides respectively. They also contained highly similar zinc finger domains. The phylogenetic analyses showed that the two isoforms and WTla of E. coioides were clustered in the same branch. The semi-quantization results revealed that high expression levels of WT1 gene can be found in gonads, kid- ney,intestine,spleen and heart. The real-time PCR results showed that WT1 gene was expressed at each stage of the gonad development, and the expression increased first and then decreased.