Identification, sequence structure and expression characteristics analysis of troponin genes of Siniperca chuatsi
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摘要: 【目的】探究鳜(Siniperca chuatsi)骨骼肌生长发育中肌钙蛋白基因的表达特征,为鳜生长发育的研究提供基础资料。【方法】基于鳜转录组数据与基因组数据联合分析,筛选鉴定出鳜肌钙蛋白基因,通过BioEdit、ClustalW、MEGA5.0、SWISS-MODEL及SMART等在线软件对鳜肌钙蛋白进行生物信息学分析,并以实时荧光定量PCR检测鳜不同肌肉组织中肌钙蛋白基因的表达特异及孵化后10~60 d(10~60 dph)骨骼肌中肌钙蛋白基因的表达规律。【结果】共鉴定出11个鳜肌钙蛋白基因[3个TnnC基因(TnnC1a、TnnC1b和TnnC2)、5个TnnI基因(TnnI1a、TnnI1b、TnnI1c、TnnI1al和TnnI2)及3个TnnT基因(TnnT1、TnnT2a和TnnT3a)],各基因亚型间在基因结构、结合位点及蛋白三级结构存在明显差异,11个鳜肌钙蛋白基因分布在4条染色体上。鳜肌钙蛋白基因具有组织特异性:TnnC1a基因在慢肌中高表达,在心肌中的相对表达量次之;TnnC1b基因在慢肌中也呈高表达,除心肌外,TnnC1a和TnnC1b基因在其他类型肌肉组织中具有相似的表达模式;TnnC2基因在快肌和颊肌中高表达;TnnI1a、TnnI1c和TnnI1al基因在慢肌中高表达,且具有相似的表达模式;TnnI1b基因在心肌中高表达,慢肌中的相对表达量次之;TnnI2基因在快肌和颊肌中高表达;TnnT1基因在慢肌中高表达,TnnT2a基因在心肌中高表达,TnnT3a基因在快肌和颊肌中高表达。随着发育时间的推移,TnnC1a、TnnC1b、TnnI1b、TnnI1c、TnnI1al和TnnT1等基因的表达显著下调(P<0.05),但TnnC2、TnnI2和TnnT3a基因在30~60 dph仍维持在相对较高的表达水平。【结论】基于鳜转录组数据与基因组数据联合分析,共鉴定出11个鳜肌钙蛋白基因,其中TnnC2、TnnI2和TnnT3a基因可作为鳜骨骼肌快肌生长发育的重要分子标记基因。Abstract: 【Objective】To explore the expression characteristics of troponin genes in the skeletal muscle growth and development of Siniperca chuatsi, to provide basic information for the study of the growth and development of S. chuatsi.【Method】Based on the joint analysis of S. chuatsi transcriptome data and genome data, troponin genes of S. chuatsi were screened and identified. Then bioinformatics analysis of the troponin was performed by online softwares such as BioEdit, ClustalW, MEGA 5.0, SWISS-MODEL and SMART. Real-time fluorescence quantitative PCR was also used to examine the expression specificity of troponin genes in different muscle tissues of S. chuatsi and their expression pattern in the skeletal muscle from 10 to 60 d(10-60 dph) after hatching. 【Result】A total of 11 troponin genes of S. chuatsi were identified: [3 TnnC genes(TnnC1a, TnnC1b and TnnC2), 5 TnnI genes(TnnI1a, TnnI1b, TnnI1c, TnnI1al and TnnI2), and 3 TnnT genes(TnnT1, TnnT2a and TnnT3a)]. They distributed on four chromosomes, with obvious differences in gene structure, binding sites and tertiary structure of protein. These genes were tissue-specific: TnnC1a gene was highly expressed in slow muscle, followed by cardiac muscle; TnnC1b gene was also highly expressed in slow muscle, and the TnnC1a and TnnC1b genes had a similar expression pattern in other types of muscle tissues except in cardiac muscle; TnnC2 gene was highly expressed in fast muscle and cheek; TnnI1a, TnnI1c and TnnI1al genes were highly expressed in slow muscle with similar expression patterns; TnnI1b gene was highly expressed in cardiac muscle and followed by slow muscle; TnnI2 gene was highly expressed in fast muscle and cheek; TnnT1 gene was highly expressed in slow muscle, the TnnT2a gene was highly expressed in cardiac muscle and TnnT3a gene was highly expressed in fast muscle and cheek.The expressions of TnnC1a, TnnC1b, TnnI1b, TnnI1c, TnnI1al and TnnT1 genes were significantly down-regulated with development time increasing(P<0.05), but the expression of TnnC2, TnnI2 and TnnT3a genes still maintained at a relatively high level at 30-60 dph. 【Conclusion】According to the joint analysis of S. chuatsi transcriptome data and genome data, a total of 11 troponin genes of S. chuatsi are identified, among which TnnC2, TnnI2 and TnnT3a genes can be used as important molecular marker genes for the growth and development of fast skeletal muscle of S. chuatsi.
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