黄颡鱼肌细胞生成素基因cDNA克隆及其在雌雄个体中的差异表达

Cloning and expression analysis of the Myo G gene in male and female yellow catfish, Pelteobagrus fulvidraco

为了探究黄颡鱼(Pelteobagrus fulvidraco)肌细胞生成素(Myogenin,MyoG)基因的组成结构、功能特性以及在雌雄个体组织中的表达差异,本研究利用RT-PCR和RACE技术克隆获得黄颡鱼MyoG基因1346 bp全长cDNA序列,其中序列包括63 bp的5′非翻译区、521 bp的3′非翻译区和762 bp的开放阅读框(ORF),ORF编码253个氨基酸。氨基酸序列包含MRFs基因家族的Basic结构域和螺旋–环–螺旋(HLH)结构域,不含信号肽,没有跨膜结构,定位于细胞核内。氨基酸序列与蓝鲇(Ictalurus furcatus)同源性高达94.1%,基于氨基酸序列构建的NJ进化树显示黄颡鱼MyoG基因与同属鲇科鱼类关系最近,表明MyoG基因在物种间比较保守。实时荧光定量PCR检测显示,MyoG基因在雌雄个体心脏、肌肉等8种组织中均有表达,但在肌肉中的表达量显著高于其他组织(P<0.05),且在雄性中的表达量显著高于雌性(P<0.05);Western blot检测MyoG蛋白在雄性肌肉中的表达量高于雌性,推测其可能是造成黄颡鱼雌雄生长差异的因素之一。本研究不仅为黄颡鱼雌雄生长差异和肌肉发育调控提供了研究基础,也为黄颡鱼快速生长新品种(系)选育提供理论参考。

Myogenin （MyoG） is a vital member of the myogenic regulatory factors （MRFs）, and plays an essential role in regulating myocyte development and maturity. Pelteobagrusfulvidraco, one of the most economically important fish in China, displays sexual dimorphism in both size and growth. To characterize the MyoG gene and its expression in male and female P fulvidraco, we cloned the full-length MyoG eDNA of this species with reverse transcription-PCR and the rapid amplification of eDNA ends. The MyoG eDNA included a 63-bp complete 5＇ untranslated region （5＇-UTR）, a 521-bp 3＇-UTR, and a 762-bp open reading frame encoding a 253-amino-acid peptide containing a basic arginine-rich domain and the helix-loop-helix structural domain typical of MRFs. The amino acid sequence was more similar to that of Ictalurus furcatus （94.1%） than to those of other species. A phylogenetic analysis based on amino acid sequences showed that P fulvidraco MyoG clustered with those of/. punctatus, 1. furcatus, and Ameiurus catus of the Siluriformes These results indicate a high degree of conservation during MyoG gene evolution. An expression analysis of MyoG in the heart, brain, liver, kidney, muscle, stomach, intestine, and gill showed that it is only significantly highly expressed in muscle tissue （P〈0.05）. Western blot analysis demonstrated that MyoG protein expression was higher in males than in females. We infer that MyoG plays an important role in the muscle development and growth of male and female P fulvidraco based on its expression characteristics at the gene and protein levels. These results extend our understanding of MyoG function, clarify the growth and differentiation of male and female P fulvidraco, and provide a reference for the selective breeding of new P. fulvidraco strains.