人喉癌基因表达相对定量研究中内参基因的验证

Validation of internal reference genes for relative quantitation studies of gene expression in human laryngeal cancer.

作者信息

Wang Xiaofeng, He Jinting, Wang Wei, Ren Ming, Gao Sujie, Zhao Guanjie, Wang Jincheng, Yang Qiwei

机构信息

Department of Stomatology, China-Japan Union Hospital, Jilin University , Changchun , China.

Department of Neurology, China-Japan Union Hospital, Jilin University , Changchun , China.

出版信息

PeerJ. 2016 Dec 8;4:e2763. doi: 10.7717/peerj.2763. eCollection 2016.

BACKGROUND

The aim of this study was to determine the expression stabilities of 12 common internal reference genes for the relative quantitation analysis of target gene expression performed by reverse transcription real-time quantitative polymerase chain reaction (RT-qPCR) in human laryngeal cancer.

METHODS

Hep-2 cells and 14 laryngeal cancer tissue samples were investigated. The expression characteristics of 12 internal reference gene candidates (18S rRNA, GAPDH, ACTB, HPRT1, RPL29, HMBS, PPIA, ALAS1, TBP, PUM1, GUSB, and B2M) were assessed by RT-qPCR. The data were analyzed by three commonly used software programs: geNorm, NormFinder, and BestKeeper.

RESULTS

The use of the combination of four internal reference genes was more appropriate than the use of a single internal reference gene. The optimal combination was PPIA + GUSB + RPL29 + HPRT1 for both the cell line and tissues; while the most appropriate combination was GUSB + RPL29 + HPRT1 + HMBS for the tissues.

CONCLUSIONS

Our recommended internal reference genes may improve the accuracy of relative quantitation analysis of target gene expression performed by the RT-qPCR method in further gene expression research on laryngeal tumors.

背景

本研究的目的是确定12种常见内参基因在人喉癌中通过逆转录实时定量聚合酶链反应（RT-qPCR）进行靶基因表达相对定量分析时的表达稳定性。

方法

对Hep-2细胞和14例喉癌组织样本进行研究。通过RT-qPCR评估12种候选内参基因（18S rRNA、GAPDH、ACTB、HPRT1、RPL29、HMBS、PPIA、ALAS1、TBP、PUM1、GUSB和B2M）的表达特征。数据通过三种常用软件程序进行分析：geNorm、NormFinder和BestKeeper。