松江鲈β-肌动蛋白基因全长cDNA的克隆和序列分析

目的]获得松江鲈β-肌动蛋白基因cDNA全长序列,并检测松江鲈β-肌动蛋白在组织中的表达。方法]以松江鲈肌肉总RNA为模板,采用RT-PCR、5′-RACE和3′-RACE的方法扩增β-肌动蛋白基因cDNA片段,用RT-PCR方法检测组织表达。结果]获得了松江鲈β-肌动蛋白基因cDNA的3个片段,测序后拼接得到1 905 bp全长cDNA序列,其包含了1 128个核苷酸的开放性阅读框,翻译编码375个氨基酸。核苷酸和氨基酸同源性分析发现,松江鲈β-肌动蛋白基因序列与点带石斑鱼、军曹鱼、红鲷鱼等同源性相对较高,与哺乳动物和鸟类同源性相对较低;系统发育分析表明,松江鲈β-肌动蛋白与点带石斑鱼关系最近。RT-PCR分析表明,该基因在检测的肌肉、肝脏、肠和脑4种组织均有表达。结论]首次得到了β-肌动蛋白基因cDNA全长序列,并证明了松江鲈的β-肌动蛋白基因非常保守。

Identification and Characterization of Beta-actin Gene in Rough-skinned Sculp,Trachidermus fasciatus

Objective] The purpose was to reveal the sequence and characteristic of Trachidermus fasciatus beta-actin gene.Method] The total RNA of muscle in Trachidermus fasciatus was isolated.Three fragments of beta-actin gene in Trachidermus fasciatus were amplified from muscle total RNA by RT-PCR,5′-RACE and 3′-RACE.Result] A full-length of 1 905 bp beta-actin cDNA sequence in Trachidermus fasciatus were obtained,which included a 1 128 bp complete open reading frame encoding a 375 amino acids peptide,a 103 bp long 5′-untranslated region,and a 674 bp long 3′-untranslated region.Sequence aligments of nucleotide and amino acid sequence revealed that Trachidermus fasciatus beta-actin has high similarity with Epinephelus coioides,Rachycentron canadum,Chrysophrys auratus beta-actin and relatively low similarity with mammalian and birds beta-actin.The phylogenetic analysis showed that Trachidermus fasciatus beta-actin has nearest relationship with Epinephelus coioides.In the four examined tissues of muscle,liver,intestine and brain,the beta-actin gene was expressed strongly in all tissues.Conclusion] The complete sequence of beta-actin gene in Trachidermus fasciatus was firstly obtained,which was highly conserved.