Reference genes selection for transcript normalization in kenaf (Hibiscus cannabinus L.) under salinity and drought stress

PeerJ. 2015 Nov 26;3:e1347. doi: 10.7717/peerj.1347. eCollection 2015.

Abstract

Kenaf (Hibiscus cannabinus) is an economic and ecological fiber crop but suffers severe losses in fiber yield and quality under the stressful conditions of excess salinity and drought. To explore the mechanisms by which kenaf responds to excess salinity and drought, gene expression was performed at the transcriptomic level using RNA-seq. Thus, it is crucial to have a suitable set of reference genes to normalize target gene expression in kenaf under different conditions using real-time quantitative reverse transcription-PCR (qRT-PCR). In this study, we selected 10 candidate reference genes from the kenaf transcriptome and assessed their expression stabilities by qRT-PCR in 14 NaCl- and PEG-treated samples using geNorm, NormFinder, and BestKeeper. The results indicated that TUBα and 18S rRNA were the optimum reference genes under conditions of excess salinity and drought in kenaf. Moreover, TUBα and 18S rRNA were used singly or in combination as reference genes to validate the expression levels of WRKY28 and WRKY32 in NaCl- and PEG-treated samples by qRT-PCR. The results further proved the reliability of the two selected reference genes. This work will benefit future studies on gene expression and lead to a better understanding of responses to excess salinity and drought in kenaf.

Keywords: Gene expression; Kenaf (Hibiscus cannabinus L.); Reference gene; Salinity and drought stress.

Grant support

This work was funded by the 948 project of the Agricultural Department of China (2013-Z70), National bast fiber germplasm resources project of China (K47NI201A) and National bast fiber research system of China (CARS-19-E06). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.