摘要
采用同源克隆和末端快速扩增(RACE)方法,克隆了半滑舌鳎中GnRHR全长cDNA。通过基因全长以及推断的氨基酸序列分析,得知该序列包含一个7tm-1保守结构域,为G蛋白偶联受体家族成员。利用PHYLIP3.5c邻位相连法以及DNAstar中的CLUSTALW方法对相应的氨基酸序列进行了聚类分析和序列相似度分析。结果表明,半滑舌鳎GnRHR与鲈鱼、琥珀鱼、虹鳟以及青鳉等鱼类中的GnRHR聚为一支,亲缘关系较近,且相似度较高,分别为90.1%、89.7%、79.0%以及78.3%,而与高等哺乳动物人、小鼠相似性仅分别为18.8%和16.2%,亲缘关系较远。应用半定量RT-PCR技术分析其组织表达,发现GnRHR广泛表达于各个组织,但表达量差异较大,在性腺、脑和肾中表达量较高,其它组织较弱。
Full-length cDNA encoding GnRHR was cloned from half smooth tongue sole Cynoglossus semilaevis by homology cloning and RACE PCR approach. According to the full length of the cDNA and deduced amino acid sequence, a 7tm-1 conservative domain was found and the cDNA was proved to be one of the G protein-coupled transmembrane family members. The rooted phylogenetic tree, analysis of identity of GnRHR, and other representative sequences were constructed by the neighbor-joining method within the package PHYLIP 3,5c and CLUSTAL W method, respectively. The results indicated that the GnRHR of the tongue sole was clustered together with those of all other fish GnRHR, for example, the similarity was 90.1%, 89.7%, 79%, 78.3% when compared with striped sea bass, amberjack, trout, and medaka, respectively. In contrast, similarity between the GnRHR of the tongue sole and those of human and mouse was much lower （18.8% and 16.2%, respectively）. Semi-quantitative RT-PCR analysis demonstrated that GnRHR was ubiquitously expressed in all tissues, although the expression level was not homogeneous. High levels of GnRHR transcript were detected in gonad, brain and kidney, and other tissues had lower levels.
出处
《海洋与湖沼》
CAS
CSCD
北大核心
2011年第4期543-548,共6页
Oceanologia Et Limnologia Sinica
基金
中央级公益性科研院所基本科研业务费专项资金
2060302号
关键词
半滑舌鳎
GnRHR
克隆
组织特异性表达
Cynoglossus semilaevis, GnRHR, Cloning, Tissue-specific expression
作者简介
李风铃，助理研究员，E-mail：milyandtt@163．com
通讯作者：王清印，研究员，博士生导师，E-mail：qywang@public．qd．sd．cn
