Exploring Valid Reference Genes for Quantitative Real-time PCR Analysis in Plutella Xylostella (Lepidoptera: Plutellidae)

Overview

Journal Int J Biol Sci

Specialty Biology

Date 2013 Aug 29

PMID 23983612

Citations 91

Authors

Wei Fu

Wen Xie

Zhuo Zhang

Shaoli Wang

Qingjun Wu

Yong Liu

Xiaomao Zhou

Xuguo Zhou

Youjun Zhang

Affiliations

Soon will be listed here.

Abstract

Quantitative real-time PCR (qRT-PCR), a primary tool in gene expression analysis, requires an appropriate normalization strategy to control for variation among samples. The best option is to compare the mRNA level of a target gene with that of reference gene(s) whose expression level is stable across various experimental conditions. In this study, expression profiles of eight candidate reference genes from the diamondback moth, Plutella xylostella, were evaluated under diverse experimental conditions. RefFinder, a web-based analysis tool, integrates four major computational programs including geNorm, Normfinder, BestKeeper, and the comparative ΔCt method to comprehensively rank the tested candidate genes. Elongation factor 1 (EF1) was the most suited reference gene for the biotic factors (development stage, tissue, and strain). In contrast, although appropriate reference gene(s) do exist for several abiotic factors (temperature, photoperiod, insecticide, and mechanical injury), we were not able to identify a single universal reference gene. Nevertheless, a suite of candidate reference genes were specifically recommended for selected experimental conditions. Our finding is the first step toward establishing a standardized qRT-PCR analysis of this agriculturally important insect pest.

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