Reference Gene Selection for Quantitative Real-time RT-PCR Normalization in the Half-smooth Tongue Sole (Cynoglossus Semilaevis) at Different Developmental Stages, in Various Tissue Types and on Exposure to Chemicals

Overview

Journal PLoS One

Specialties General Medicine
Science

Date 2014 Mar 27

PMID 24667563

Citations 13

Authors

Conghui Liu

Nian Xin

Yi Zhai

Liming Jiang

Jieming Zhai

Quanqi Zhang

Jie Qi

Affiliations

Soon will be listed here.

Abstract

Quantitative real time RT-PCR has been described as the most sensitive method for the detection of low abundance mRNA. To date, no reference genes have been screened in the half-smooth tongue sole (Cynoglossus semilaevis). The aim of this study was to select the most stable genes for quantitative real-time RT-PCR. Eight housekeeping genes (18S, TUBA, B2M, ACTB, EF1A, GAPDH, RPL17 and UBCE) were tested at different developmental stages, in different tissues, and following exposure to the drug SB-431542. Using geNorm, BestKeeper and NormFinder software, GAPDH/B2M, GAPDH/18S and UBCE/GAPDH were identified as the most suitable genes from samples taken of different developmental stages while 18S/RPL17 were consistently ranked as the best reference genes for different tissue types. Furthermore, TUBA/B2M, TUBA/UBCE and B2M/TUBA were found to be the most suitable genes in samples treated with the drug, SB-431542 by geNorm, BestKeeper and NormFinder respectively. Across both different developmental stages and tissue types, the combination of 18S and GAPDH was the most stable reference gene analyzed by Ref-Finder. To test and verify the screened reference genes, the expression profiles of LEFTY-normalized to the combination of GAPDH/18S and ACTB were presented. These results will be useful for future gene-expression studies in the half-smooth tongue sole.

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