Journal
INTERNATIONAL JOURNAL OF BIOLOGICAL SCIENCES
Volume 9, Issue 8, Pages 792-802Publisher
IVYSPRING INT PUBL
DOI: 10.7150/ijbs.5862
Keywords
Plutella xylostella; reference gene; qRT-PCR analysis; biotic factor; abiotic factor
Categories
Funding
- National Natural Science Foundation of China [31071709, 31171876]
- 863 Program [2012AA101502]
- Special Fund for Agro-scientific Research in the Public Interest [201103021]
- Beijing Key Laboratory for Pest Control and Sustainable Cultivation of Vegetables
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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 Delta Ct 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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