Identification of appropriate reference genes for RT-qPCR analysis in Juglans regia L

Li Zhou; Jianxin Niu; Shaowen Quan

doi:10.1371/journal.pone.0209424

Identification of appropriate reference genes for RT-qPCR analysis in Juglans regia L

PLoS One. 2018 Dec 18;13(12):e0209424. doi: 10.1371/journal.pone.0209424. eCollection 2018.

Authors

Li Zhou^{1

2}, Jianxin Niu^{1

2}, Shaowen Quan^{1

2}

Affiliations

¹ Department of Horticulture, College of Agriculture, Shihezi University, Shihezi, Xinjiang, P.R. China.
² Xinjiang Production and Construction Corps Key Laboratory of Special Fruits and Vegetables Cultivation Physiology and Germplasm Resources Utilization, Shihezi, Xinjiang, P.R. China.

Abstract

Reverse transcription quantitative real-time PCR (RT-qPCR) is a popular adopted technique to detect gene expression, and the selection of appropriate reference genes is crucial for data normalization. In the present study, seven candidate reference genes were screened to evaluate their expression stability in various flower buds, leaf buds, tissues and cultivars of the English walnut (Juglans regia L.) based on four algorithms (geNorm, Normfinder, Bestkeeper and RefFinder). The results demonstrated that TUA, EF1 and TUB were appropriate reference genes for flower buds at different stages of female flower buds differentiation; TUB and 18S rRNA were best for leaf buds at different stages of female flower buds differentiation; TUB and TUA were suitable for different cultivars; and ACT2, 18S rRNA and GAPDH were useful for different tissues. Moreover, the expression of ACT was not stable among different flower buds, leaf buds and cultivars. The stability of reference genes were confirmed through the analysis of the expression of SPL18 gene. These results will contribute to a reliable normalization of gene expression in J. regia.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Gene Expression Profiling / methods
Gene Expression Profiling / standards*
Genes, Plant / genetics*
Juglans / genetics*
Reference Standards
Reverse Transcriptase Polymerase Chain Reaction / standards*

Grants and funding

The study was supported by the National Science and Technology Specific projects of Xinjiang (No. 201130102-1-4) and the National Natural Science Foundation of China (No. 30560090). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.