革胡子鲶MHC基因表达qPCR分析的引物设计与评估

天津农学院学报 ›› 2016, Vol. 23 ›› Issue (4): 33-37.

革胡子鲶MHC基因表达qPCR分析的引物设计与评估

李转转¹, 王妍², 高金伟¹, 王晓梅^{1,通信作者}, 陈成勋¹, 李涛³, 莫宝霖¹

1. 天津农学院水产学院,天津市水产生态及养殖重点实验室,天津 300384;
2. 天津海友佳音生物科技股份有限公司,天津 300350;
3. 天津农学院基础科学学院,天津 300384

收稿日期:2016-03-23 出版日期:2016-12-20 发布日期:2019-10-14
通讯作者: 王晓梅（1962-）,女,天津市人,教授,博士,主要从事水产动物分子生物学的研究。E-mail：xiaomeiw@tjau.edu.cn。
作者简介:李转转（1992-）,女,山西吕梁人,本科在读,主要从事水产动物分子生物学的研究。E-mail：1297036725@qq.com。
基金资助:
天津市应用基础与前沿技术研究计划（重点项目）“革胡子鲶抗菌肽提取与鉴别及非特异免疫因子的分子解析”（15JCZDJC33500）; 天津农学院中青年骨干创新人才培养计划“渔业资源利用与生态修复”（无编号）

Design and Evaluation of Primers for Analysis on MHC Gene Expression Profiles of Clarias gariepinus by Using Real-Time Quantitative PCR Technique

LI Zhuan-zhuan¹, WANG Yan², GAO Jin-wei¹, WANG Xiao-mei¹, CHEN Cheng-xun¹, LI Tao³, MO Bao-lin¹

1. Tianjin Key Laboratory of Aqua-Ecology and Aquaculture, College of Fisheries, Tianjin Agricultural University, Tianjin 300384, China;
2. Tianjin Ocean Pal Carol Biotech CO, Ltd, Tianjin 300350, China;
3. College of Basic Science, Tianjin Agricultural University, Tianjin 300384, China

Received:2016-03-23 Online:2016-12-20 Published:2019-10-14

摘要/Abstract

摘要： 为应用实时荧光定量PCR（qPCR）技术分析革胡子鲶主要组织相容性复合体（major histocompatibility complex,MHC）基因的表达谱,本研究利用Primer 5软件设计扩增革胡子鲶MHC基因片段的引物并进行qPCR评估。结果显示：引物MHCF₁-MHCR₃在qPCR扩增时,融解曲线为单一尖锐峰;标准曲线分析显示：引物的扩增效率为99.3%,R²值为0.998。上述结果说明该对引物具有良好的扩增特异性和扩增效率,满足了qPCR扩增的要求。本研究为革胡子鲶MHCⅠ基因表达的qPCR分析奠定了基础。

关键词: 革胡子鲶, 主要组织相容性复合体, 实时荧光定量PCR, 融解曲线, 扩增效率

Abstract: In order to analyze the MHC gene expression profiles in Clarias gariepinus by using real-time quantitative PCR（qPCR）technique, primer pairs for amplifying the MHC gene fragments of C. gariepinus were designed using Primer 5.0 software and then assessed by means of general PCR and qPCR amplification, respectively. The results of qPCR evaluation showed that melt curve of the primer pair named MHCF₁-MHCR₃ presented a single sharp peak, and standard curve displayed that its amplification efficiency was 99.3% and R² value was 0.998. The results indicated that the primer pair had good specificity and efficiency of amplification as well as met the criteria of qPCR amplification. This work laid a foundation for analyzing MHC gene expression of C. gariepinus by using qPCR technique.

Key words: Clarias gariepinus, major histocompatibility complex, real-time fluorescence quantitative PCR, melt peak, amplification efficiency

中图分类号:

S965.128

李转转, 王妍, 高金伟, 王晓梅, 陈成勋, 李涛, 莫宝霖. 革胡子鲶MHC基因表达qPCR分析的引物设计与评估[J]. 天津农学院学报, 2016, 23(4): 33-37.

LI Zhuan-zhuan, WANG Yan, GAO Jin-wei, WANG Xiao-mei, CHEN Cheng-xun, LI Tao, MO Bao-lin. Design and Evaluation of Primers for Analysis on MHC Gene Expression Profiles of Clarias gariepinus by Using Real-Time Quantitative PCR Technique[J]. Journal of Tianjin Agricultural University, 2016, 23(4): 33-37.

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http://xuebao.tjau.edu.cn/CN/Y2016/V23/I4/33

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