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Selection of internal reference gene for normalization of reverse transcription-quantitative polymerase chain reaction analysis in Mycoplasma hyopneumoniae

文献类型：期刊论文

作者Li, Shiyang5,6; Zhou, Yanqing3,4,5; Yuan, Ting5; Feng, Zhixin4,5; Zhang, Zhenzhen5; Wu, Yuzi5; Xie, Qingyun5; Wang, Jia5; Li, Quan2; Deng, Zhibang6
刊名FRONTIERS IN VETERINARY SCIENCE
出版日期2022-07-22
卷号9页码:11
关键词Mycoplasma hyopneumoniae RT-qPCR real-time quantitative polymerase chain reaction internal reference genes gatB virulence
DOI10.3389/fvets.2022.934907
通讯作者Yu, Yanfei(yuyanfeihaha@163.com) ; Yuan, Xiaomin(187027211@qq.com)
英文摘要Mycoplasma hyopneumoniae is the etiological agent of swine enzootic pneumonia (EP), which resulting in considerable economic losses in pig farming globally. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) is a major tool for gene expression studies. However, no internal reference genes for normalization of RT-qPCR data of M. hyopneumoniae have been reported. The aim of this study was to screen the most stable genes for RT-qPCR analysis in M. hyopneumoniae under different conditions. Therefore, a total of 13 candidate internal reference genes (rpoC, Lipo, sgaB, oppB, hypo621, oppF, gyrB, uvrA, P146, prfA, proS, gatB, and hypo499) of M. hyopneumoniae filtered according to the reported quantitative proteomic analysis and the 16S rRNA internal reference gene frequently used in other bacteria were selected for RT-qPCR analysis. The mRNAs from different virulence strains (168, 168 L, J, NJ, and LH) at five different growth phases were extracted. The corresponding cycle threshold (Ct) values of the 25 reverse transcribed cDNAs using the 14 candidate genes were determined. Different internal reference genes or combinations were then screened for expression stability analysis using various statistical tools and algorithms, including geNorm, BestKeeper, and NormFinder software, to ensure the reliability of the analysis. Through further comprehensive evaluation of the RefFinder software, it is concluded that the gatB gene was the most suitable internal reference gene for samples of the different virulence strains in different growth phases for M. hyopneumoniae, followed by prfA, hypo499, and gyrB.
WOS关键词PCR
资助项目Programs of Jiangsu Agricultural Science and Technology Innovation Fund[CX (20) 3090] ; Programs of National Natural Science Foundation of China[32172860] ; Programs of National Natural Science Foundation of China[32102675] ; Programs of National Natural Science Foundation of China[31900159] ; Natural Science Foundation of Jiangsu Province[BK20190269]
WOS研究方向Veterinary Sciences
语种英语
出版者FRONTIERS MEDIA SA
WOS记录号WOS:000837217700001
源URL[http://119.78.100.183/handle/2S10ELR8/301893]  
专题中国科学院上海药物研究所
通讯作者Yu, Yanfei; Yuan, Xiaomin
作者单位1.Jiangsu Univ, Sch Food & Biol Engn, Zhenjiang, Peoples R China
2.Yangzhou Univ, Coll Vet Med, Yangzhou, Peoples R China
3.Chinese Acad Sci, Zhongshan Inst Drug Discovery, Shanghai Inst Mat Med, Zhongshan, Peoples R China
4.Nanjing Agr Univ, Coll Vet Med, Nanjing, Peoples R China
5.Minist Agr & Rural Affairs, Jiangsu Acad Agr Sci, Inst Vet Med, Key Lab Vet Biol Engn & Technol, Nanjing, Peoples R China
6.Hunan Agr Univ, Coll Vet Med, Changsha, Peoples R China
推荐引用方式
GB/T 7714		 Li, Shiyang,Zhou, Yanqing,Yuan, Ting,et al. Selection of internal reference gene for normalization of reverse transcription-quantitative polymerase chain reaction analysis in Mycoplasma hyopneumoniae[J]. FRONTIERS IN VETERINARY SCIENCE,2022,9:11.
APA Li, Shiyang.,Zhou, Yanqing.,Yuan, Ting.,Feng, Zhixin.,Zhang, Zhenzhen.,...&Yuan, Xiaomin.(2022).Selection of internal reference gene for normalization of reverse transcription-quantitative polymerase chain reaction analysis in Mycoplasma hyopneumoniae.FRONTIERS IN VETERINARY SCIENCE,9,11.
MLA Li, Shiyang,et al."Selection of internal reference gene for normalization of reverse transcription-quantitative polymerase chain reaction analysis in Mycoplasma hyopneumoniae".FRONTIERS IN VETERINARY SCIENCE 9(2022):11.

入库方式： OAI收割

来源：上海药物研究所

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