Suitable Reference Genes for Accurate Gene Expression Analysis in Parsley (Petroselinum crispum) for Abiotic Stresses and Hormone Stimuli.

Meng-Yao Li,Ai-Sheng Xiong,Feng Wang,Xiong Song

doi:10.3389/fpls.2016.01481

Abstract

Parsley, one of the most important vegetables in the Apiaceae family, is widely used in the food, medicinal, and cosmetic industries. Recent studies on parsley mainly focus on its chemical composition, and further research involving the analysis of the plant's gene functions and expressions is required. qPCR is a powerful method for detecting very low quantities of target transcript levels and is widely used to study gene expression. To ensure the accuracy of results, a suitable reference gene is necessary for expression normalization. In this study, four software, namely geNorm, NormFinder, BestKeeper, and RefFinder were used to evaluate the expression stabilities of eight candidate reference genes of parsley (GAPDH, ACTIN, eIF-4α, SAND, UBC, TIP41, EF-1α, and TUB) under various conditions, including abiotic stresses (heat, cold, salt, and drought) and hormone stimuli treatments (GA, SA, MeJA, and ABA). Results showed that EF-1α and TUB were the most stable genes for abiotic stresses, whereas EF-1α, GAPDH, and TUB were the top three choices for hormone stimuli treatments. Moreover, EF-1α and TUB were the most stable reference genes among all tested samples, and UBC was the least stable one. Expression analysis of PcDREB1 and PcDREB2 further verified that the selected stable reference genes were suitable for gene expression normalization. This study can guide the selection of suitable reference genes in gene expression in parsley.

Highlights

Gene expression analysis is an important tool from studying the complex biological processes, such as signal transduction, metabolic pathways, and plant development
The eight candidate reference genes (GAPDH, ACTIN, eIF4α, SAND, UBC, Tap42-interacting protein of 41 kDa gene (TIP41), Elongation factor1α gene (EF-1α), and TUB) were selected from the transcriptome database, and they were chosen as their corresponding homologs genes performed well in quantitative real-time reverse transcription PCR (qPCR)
QPCR is one of the most sensitive methods that can detect the low expression of target genes (Bustin, 2000)

Summary

Introduction

Gene expression analysis is an important tool from studying the complex biological processes, such as signal transduction, metabolic pathways, and plant development. In the qualitative or quantitative analysis of the expression of the sample or target gene, qPCR is the most effective method because of its simplicity, high sensitivity and specificity (Wong and Medrano, 2005). Selection and Validation of Reference Genes in Parsley. To date, universal reference gene in plants or animals has not been reported yet (Warrington et al, 2000; Schmittgen and Zakrajsek, 2000). Just borrowed the reference genes in other species might get the wrong expression model. Evaluation of the stability of reference genes is important

Methods

Results

Conclusion