泰国斗鱼抗缪勒氏管激素(AMH)基因cDNA克隆及表达

Molecular Cloning and Tissue Expression of Thailand Betta(Betta splendens) Anti-Müllerian Hormone(AMH)

利用RACE(Rapid-amplification of cDNA ends)克隆技术从泰国斗鱼(Betta splendens)精巢中克隆抗缪勒氏管激素(Anti-Müllerian hormone,AMH)基因的c DNA序列,分析该基因序列与其他物种的差异,并用RT-PCR半定量方法分析其组织表达的性别差异。结果表明:泰国斗鱼AMH基因的cDNA序列全长为1 804 bp,其中开放读码框为1 596 bp,编码532个氨基酸残基,与其他物种的氨基酸序列差异较大,同源性最高仅55%;AMH前体蛋白的系统进化树显示,泰国斗鱼与尖吻鲈(Lates calcarifer)亲缘关系最近;AMH基因的组织表达有明显的组织特异性及性别差异,在性腺中表达量最高,雄性个体脑、脾和肌肉次之,垂体、肝、肾、心和肠中未检测到AMH表达,雌性个体脾、心脏和肌肉次之,脑、垂、肝和肾中未检测AMH表达。

The Anti-Müllerian hormone c DNA of the Thailand betta（Betta splendens） was isolated from the testis by rapid-amplification of c DNA ends（RACE）. Clastal alignment was used to analyse the sequence divergence of AMH between Thailand betta and other species by studying alignment and phylogenetic trees and the tissue expression of AMH was performed using semi-quantitative RT-PCR. The results showed that the Thailand betta AMH c DNA sequence consisted of 1804 nucleotides with an open reading frame（ORF） of 1 596 bp. The deduced AMH consisted of 532 amino-acid residues. Sequence comparison revealed that the Thailand betta AMH shared only 55% similarity to the AMH of Lates calcarifer. Based on the results from phylogenetic tree analysis, the Thailand betta AMH was more closely related to the AMH of Lates calcarifer. In addition, Thailand betta AMH displayed tissue and sex specific expression patterns in the gonad. In the male fish, strong expression of AMH was observed in the testis but only moderate expression was found in the brain, spleen and muscle. No expression signal was detected in the pituitary, liver, kidney, heart and intestine. In the females, high expression level of Thailand betta AMH was observed in the ovary and only moderate m RNA level was found in the spleen, heart and muscle. No expression of AMH was detected in the brain, pituitary, liver and spleen.