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罗双双,高泽霞,冯兵,张曼曼,曹文怡,周小云.二倍体和四倍体泥鳅qRT-PCR分析中内参基因优选[J].华中农业大学学报,2019,38(5):1-8
二倍体和四倍体泥鳅qRT-PCR分析中内参基因优选
Screening of optimal internal reference genes for qRT-PCR in diploid and tetraploid loach, Misgurnus anguillicaudatus
投稿时间:2019-05-05  
DOI:
中文关键词:  泥鳅  二倍体  四倍体  多倍体鱼类  表型变异  基因表达变异  qRT-PCR  内参基因  稳定表达
英文关键词:Misgurnus anguillicaudatus  diploid  tetraploid  polyploid fish  phenotypic variation  gene expression variation  qRT-PCR  internal control genes  stable expression
基金项目:国家重点研发计划“蓝色粮仓科技创新”重点专项(2018YFD0900205)
作者单位E-mail
罗双双 华中农业大学水产学院/农业动物遗传育种与繁育教育部重点实验室武汉 430070 598874009@qq.com 
高泽霞 华中农业大学水产学院/农业动物遗传育种与繁育教育部重点实验室武汉 430070  
冯兵 华中农业大学水产学院/农业动物遗传育种与繁育教育部重点实验室武汉 430070  
张曼曼 华中农业大学水产学院/农业动物遗传育种与繁育教育部重点实验室武汉 430070  
曹文怡 华中农业大学水产学院/农业动物遗传育种与繁育教育部重点实验室武汉 430070  
周小云 华中农业大学水产学院/农业动物遗传育种与繁育教育部重点实验室武汉 430070 zhouxy@mail.hzau.edu.cn 
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中文摘要:
      为筛选在泥鳅中稳定表达的内参基因,以确保泥鳅基因表达分析结果的可靠性,选取ACTB、TUBA、EF1A、GAPDH、TUBB和18S rRNA 6个常用的内参基因作为候选内参基因,分别用BestKeeper、geNorm、NormFinder、ΔCt和RefFinfer软件分析这6个基因在二倍体和四倍体泥鳅胚胎发育阶段、成鱼组织间以及倍性间表达的稳定性。结果显示,在胚胎发育各阶段,TUBB和TUBA的表达最稳定,可作为胚胎发育阶段qRT-PCR分析的内参基因;其中,TUBB在泥鳅倍性间的表达差异不显著,因此,可作为泥鳅胚胎发育中跨倍性qRT-PCR研究的内参基因。在成鱼各组织中,ACTB和18S rRNA的表达最稳定,可用作组织间qRT-PCR分析的内参基因;其中,18S rRNA在倍性间的组织表达差异不显著,因此,可作为泥鳅组织间跨倍性qRT-PCR的内参基因。
英文摘要:
      To identify the optimal internal reference genes for qRT-PCR analysis in loach (Misgurnus anguillicaudatus), six commonly used reference genes, including ACTB, TUBA, EF1A, GAPDH,TUBB and 18S rRNA were selected as candidates, and their expression stability during different developmental stages, across tissues and between ploidy were evaluated by using BestKeeper, geNorm, NormFinder, ΔCt and RefFinfer software. The results showed that, during the process of embryonic development, the expression levels of TUBB and TUBA were more stable than the other genes, suggesting that they can be used as reference genes. Besides, the expression level of TUBB showed no significant difference between diploids and tetraploids (P>0.05), suggesting it is the most suitable internal reference genes for qRT-PCR analysis during embryonic development between ploidy. Among different tissues of diploids and tetraploids, the expression levels of ACTB and 18S rRNA were more stable than the other genes, thus they can be used as internal control genes. Of which,18S rRNA was considered as the most suitable internal control genes, because the expression level of it exhibited no significant difference in different tissues between diploids and tetraploids (P>0.05).
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