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2025年5月7日 星期三
  2015, Vol. 23 Issue (3): 359-368    
  研究论文与报告 本期目录 | 过刊浏览 | 高级检索 |
鹅肌球蛋白重链1基因MyHC1全长cDNA克隆、序列及胚胎期表达特征分析
李艳艳1,1,邵芳1,1,吴小娟1,1,石亦静1,1,郁建锋1,1,顾志良2
1.
2. 常熟理工学院
Molecular Cloning, Sequence and Expression Characterization of Myosin Heavy Chain 1 Gene (MyHC1) in Goose (Anser anser) at Embryonic Period
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摘要 肌球蛋白是组成肌原纤维粗丝的主要成分,在肌肉生长发育和运动收缩过程起重要作用。本研究采用反转录PCR和快速扩增cDNA末端(rapid amplification cDNA ends, RACE)方法,从太湖鹅(Anser anser)肌肉中克隆到肌球蛋白重链1基因(myosin heavy chain 1, MyHC1)的全长cDNA,并运用生物信息学的方法对其进行分析,实时荧光定量PCR(Real-time fluorescent quantitative PCR, qRT-PCR)检测MyHC1基因在鹅胚胎期的表达情况。研究结果表明,鹅MyHC1基因(GenBank 登录号: KM675469)cDNA全长6 028 bp,包含5 823 bp的开放性阅读框,57 bp的5'端非编码区和148 bp的3'端非编码区,共编码1 940个氨基酸。经预测,鹅MyHC1基因编码的蛋白质由31 478个原子组成,分子式为C9746H15817N2753O3096S66,相对分子质量为223 kD,等电点为5.62,平均亲水性为-0.786,属于不稳定亲水蛋白。在线预测鹅和鸡(Gallus gallus)的MyHC1蛋白质三维结构呈高度相似,由多螺旋和折叠片聚集成球状头部,并带有长纤维状α-螺旋尾部。鹅MyHC1的编码区(coding sequence, CDS)序列与鸡的同源性最高,为92.86%,与哺乳类同源性多数在84%左右,与两栖类同源性相对较低。鹅与鸡氨基酸同源性最高,为96.45%,与哺乳类的同源性多数在90%左右,与非洲爪蛙(Xenopus laevis)同源性较低,为78.13%。系统进化树分析表明,哺乳动物形成一个大分支,两栖类自成一支,鹅和红原鸡聚成一支,分子进化地位的关系最近,验证了鹅和鸡属于近缘物种。qRT-PCR检测到MyHC1基因在胚胎期第7 天开始表达,以后表达量逐渐升高,在15 d表达量达到高峰后下降,25 d之后逐渐趋于平稳,表明,MyHC1基因在鹅胚胎期mRNA水平的表达量呈先上升再下降的趋势。本研究首次获得了鹅MyHC1的全长cDNA序列、分子的结构特点和表达特征,显示该基因在鹅肌肉生长发育过程起重要作用,为该基因的功能及鹅肉质性状研究提供了基础资料。
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李艳艳
邵芳
吴小娟
石亦静
郁建锋
顾志良
关键词 肌球蛋白重链1(MyHC1)胚胎期组织表达分析    
Abstract:Myosin is the major component of myofibrillar thick wire, and plays important roles in the growth and muscle contraction. In this study, reverse transcription PCR and rapid amplification cDNA ends (RACE) methods were used to obtain the full-length cDNA of myosin heavy chain1 gene (MyHC1) in Anser anser. The bioinformatic analysis and expression patter of MyHC1 mRNA at embryonic period detected by Real-time fluorescent quantitative PCR (qRT-PCR) were performed, respectively. The results showed that the MyHC1(GenBank No. KM675469) cDNA full-length was of 6 028 bp including a 5 823 bp open reading frame, 57 bp 5'UTR and 148 bp 3'UTR, and encoded a polypeptide of 1 940 amino acids. The MyHC1 protein was predicted to compose of 31 478 atoms with the formula of C9746H15817N2753O3096S66 and a kind of unstable and hydrophilic protein, with a calculated relative molecular mass of 223 kD, pI of 5.62 and an average hydropathicity of -0.786. The 3-dimensional structure of MyHC1 protein prediction online between Anser anser and Gallus gallus was highly similar, many spiral and folded pieces gathered into globular head with a long fibrous alpha helix tail. Multiple sequence comparisons showed that Anser anser MyHC1 had high homology with Gallus gallus at 92.86%, about 84% with most mammals, and relatively low with amphibians. Further amino acid sequence comparisons showed high homology with Gallus gallus at 96.45%, about 90% with most mammals, and relatively low with Xenopus laevis at 78.13%. Then phylogenetic relationship analysis indicated that mammals formed a large branch, and amphibians formed another branch. Anser anser clustered with Gallus gallus which was similar to the results of the biological classification, which verified they belonged to related species. qRT-PCR result showed that MyHC1 expressed form the early 7 d, increased until 15 d, reached a peak later, then decreased and gradually stabilized after 25 d, showing MyHC1 mRNA expression level of a downward trend after the first rise. This study provides the full-length cDNA sequence and characteristics of geese MyHC1 gene and protein, suggesting that it plays an important role in the process of muscle growth which sipplies the basic information for its function and goose meat quality traits research.
Key wordsAnser anser    Myosin heavy chain 1(MyHC1)    Embryonic stage    Tissue expressional pattern
收稿日期: 2014-10-08      出版日期: 2015-02-02
基金资助:江苏省六大人才高峰项目;苏州市科技支撑计划
通讯作者: 顾志良     E-mail: zhilianggu88@hotmail.com
引用本文:   
李艳艳 邵芳 吴小娟 石亦静 郁建锋 顾志良. 鹅肌球蛋白重链1基因MyHC1全长cDNA克隆、序列及胚胎期表达特征分析[J]. , 2015, 23(3): 359-368.
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http://journal05.magtech.org.cn/Jwk_ny/CN/     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2015/V23/I3/359
 
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