尼罗罗非鱼Dmrt基因的序列分析

Sequence Analysis of Dmrt Genes in Oreochromis niloticus

参照不同物种DM结构域设计了1对兼并引物,扩增出1条带,长约150 bp。对扩增产物进行克隆,阳性克隆采用SSCP分析筛选不同基因片段,进一步对筛选的不同基因进行了DNA测序。结果获得2个不同的Dmrt基因,采用BLAST方法与人类相应Dmrt基因进行序列比对,发现其与人类相应的Dmrt基因的氨基酸序列一致性分别为89.1%和100.0%。根据尼罗罗非鱼(Oreochromis niloticus)的拉丁文名,按习惯将其命名为OnDmrt1和OnDmrt2。根据扩增出的Dmrt1DM-domain的序列设计上游引物,用3′RACE(rapid amplification of cDNA ends)法分离和测定了Dmrt1cDNA的3′序列,得到1108 bp的片段,共编码262个氨基酸,与奥利亚罗非鱼、虹鳟、新月鱼、青鳉等动物的Dmrt1的氨基酸序列进行比较,同源性分别为99%、62%、73%和41%。

A 150 bp-long Dmrt gene of Oreochromis niloticus were amplified using a pair of degenerated primers which were designed based on the conservative DM domain of different species. Single strand confirmation polymorphism analysis was conducted on the positive clones to select gene fragments. Two different Dmrt genes were obtained and sequenced respectively. The two Dmrt genes were named as OnDmrtl and OnDmrt2 according to their high homologies （89. 1% and 100. 0%, respectively） to corresponding Human Dmrt genes. A rapid amplification of cDNA ends （RACE） was used for isolation of the length cDNA of Dmrt1 from the testis of O. niloticus. A fragment about 1 108 bp by 3＇RACEPCR was amplified, encoding 262 amino acids. Its similarities with 0. aurea, Oncorhynchus mykiss, Xiphophorus maculatus and Oryzias latipes, were 99%, 62%, 41%, 73%, respectively. Theses results are helpful for the studies on sex control mechanism of Dmrtl in O. niloticus.