Selection of Internal References for RT-qPCR Assays in Neurofibromatosis Type 1 (NF1) Related Schwann Cell Lines

Overview

Journal PLoS One

Specialties General Medicine
Science

Date 2021 Feb 25

PMID 33630851

Citations 3

Authors

Yi-Hui Gu

Xi-Wei Cui

Jie-Yi Ren

Man-Mei Long

Wei Wang

Cheng-Jiang Wei

Rehanguli Aimaier

Yue-Hua Li

Man-Hon Chung

Bin Gu

Qing-Feng Li

Zhi-Chao Wang

Affiliations

Soon will be listed here.

Abstract

Real-time quantitative PCR (RT-qPCR) has been widely applied in uncovering disease mechanisms and screening potential biomarkers. Internal reference gene selection determines the accuracy and reproducibility of data analyses. The aim of this study was to identify the optimal reference genes for the relative quantitative analysis of RT-qPCR in fourteen NF1 related cell lines, including non-tumor, benign and malignant Schwann cell lines. The expression characteristics of eleven candidate reference genes (RPS18, ACTB, B2M, GAPDH, PPIA, HPRT1, TBP, UBC, RPLP0, TFRC and RPL32) were screened and analyzed by four software programs: geNorm, NormFinder, BestKeeper and RefFinder. Results showed that GAPDH, the most frequently used internal reference gene, was significantly unstable between various cell lines. The combinational use of two reference genes (PPIA and TBP) was optimal in malignant Schwann cell lines and the use of single reference genes (PPIA or PRLP0) alone or in combination was optimal in benign Schwann cell lines. These recommended internal reference gene selections may improve the accuracy and reproducibility of RT-qPCR in gene expression analyses of NF1 related tumors.

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