Abstract
qRT-PCR is one of the most popular approaches to analyze specific gene expression level, and stably expressed reference genes are essential to obtain reliable results. However, many reference genes are only stable under certain circumstances and different reference genes might be required in different experiments. High temperature is a common stress that affects rice endosperm development and it has become a hot topic recently. Although study about reference genes at different developmental stages in rice has been reported, these genes may not be suitable to study high temperature mediated responses especially in endosperm. In our quest for proper reference genes to quantify gene expression in rice endosperm under high temperature, we studied 6 candidate genes selected from the transcriptome data and 11 housekeeping genes. All genes were analyzed with qRT-PCR and the expression stability was assessed with software geNorm and NormFinder. Fb15 and eIF-4a were identified as the two most stable genes in endosperm at different developmental stages, while high temperature treatment has a least effect on expression of Fb15 and UBQ5 in rice endosperm. Our results provide some good candidate reference genes for qRT-PCR normalization in rice endosperm under different temperatures.
Highlights
Quantitative reverse transcription polymerase chain reaction is an efficient, costeffective and reproducible method for quantifying gene transcript levels in molecular biology research [1]. qRT-PCR is widely used to analyze gene expression in plant tissues at different developmental stages and/or under different biotic or abiotic stresses [2,3,4]
Since we focused on the durable effects of high temperature on endosperm development, RNA samples were collected 5 days later for RNA-seq
A lower coefficient of variation (CV) indicates less variation, 712 genes with lowest CV (CV
Summary
Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is an efficient, costeffective and reproducible method for quantifying gene transcript levels in molecular biology research [1]. qRT-PCR is widely used to analyze gene expression in plant tissues at different developmental stages and/or under different biotic or abiotic stresses [2,3,4]. The accuracy of qRT-PCR is determined by a number of factors, such as mRNA quality, reverse
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