半滑舌鳎促性腺激素α亚基cDNA的克隆及组织表达特征

Cloning and mRNA expression pattern of common glycoprotein α subunit of GTH in tongue sole Cynoglossus semilaevis Günther

关键词:  半滑舌鳎  促性腺激素α亚基基因  cDNA克隆  qRT-PCR

Key words:  Cynoglossus semilaevis Günther  Common glycoprotein α subunit gene of GTH  cDNA cloning  Quantitative Real-Time RT-PCR

摘要:

采用同源克隆和末端快速扩增（RACE）方法，从半滑舌鳎脑垂体中克隆了促性腺激素α亚基（CGα）全长cDNA（GenBank序列登录号：JQ364953）。半滑舌鳎CGα基因全长685bp，其开放阅读框384bp，编码了含127个氨基酸的蛋白，N端33个氨基酸为信号肽。半滑舌鳎CGα成熟肽与其他脊椎动物CGα成熟肽结构特征相似，具有10个半胱氨酸残基和两个N-糖基化位点。CGα成熟肽序列分析表明，半滑舌鳎CGα与鲽形目和鲈形目鱼类CGα同源性为60%~70%，与鲤形目鱼类CGα同源性为55%~60%。实时荧光定量RT-PCR结果表明，半滑舌鳎CGα mRNA在被检测的12个组织中均有表达，除头肾和脾脏、脾脏和肝脏间表达量差异不显著外，其他组织间表达量差异显著（P<0.05）；CGα mRNA在垂体组织中大量表达，其次是鳃、肾脏、肌肉、卵巢和脑组织，而在心脏、头肾、肝和脾等组织中表达量很低。本研究为进一步探讨促性腺激素在半滑舌鳎繁殖生理中的作用机制奠定基础。

Abstract:

Full-length cDNA encoding common glycoprotein α subunit (CGα) was firstly cloned from half smooth tongue sole Cynoglossus semilaevis Günther by means of homology cloning and RACE PCR analyses. This sequence was submitted to GenBank with accession No. JQ364953. The whole cDNA length of tongue sole CGα gene was 685bp, including a 384bp open reading frame (ORF). The predicted mature tongue sole CGα consisted of 127 amino acids preceded by a signal peptide of 33 residues. The structure of mature CGα in tongue sole CGα was similar with those in the vertebrate CGα, including 10 conserved Cysteine residues and 2 N-linked glycosylation sites. Sequence comparison showed that tongue sole CGα mature peptide had higher identity to their orthologs of percomorphs (perciforms and pleuronectiforms; 60%~70% identity) than to cypriniformes (55%~60% identity). The CG(mRNA expression of tongue sole was detected in all the examined tissues by quantitative Real-Time PCR (qRT-PCR) method. The significant test analysis (P<0.05) for expression levels of tissues revealed that head kidney and spleen as well as spleen and liver did not differ significantly from each other. But other tissues showed significant different from each other. The highest expression level was in pituitary, followed by gill, kidney, muscle, ovary and brain; and lower level expression was found in other tissues including heart, head kidney, liver and spleen.