Stable Reference Gene Selection for qRT-PCR Normalization in Strawberry (Fragaria × ananassa) Leaves under Different Stress and Light-Quality Conditions

Yuntian Ye,Yang Lu,Guangyi Wang,Yongqiang Liu,Yunting Zhang,Haoru Tang

doi:10.3390/horticulturae7110452

Abstract

Selecting an appropriate reference gene is of crucial importance for improving the accuracy of qRT-PCR analyses. In this study, strawberry (Fragaria ananassa) seedlings were subjected to different environmental conditions including heat, cold, drought, salt, white-light, blue-light, and red-light treatments. The expression levels of seven candidate reference genes, including Fa18S, FaGAPDH, FaPIRUV, FaDBP, FaHISTH4, FaACTIN1, and FaACTIN2, in the strawberry leaves were measured by qRT-PCR. Then, four programs (geNorm, NormFinder, BestKeeper, and RefFinder) were employed as tools to evaluate the expression stability of the candidate reference genes. The results showed that the expression stability of the reference genes varied under different conditions. For the cold stress and white-light treatments, FaACTIN2 was evaluated to be the most stable reference gene. FaGAPDH should be used as the reference gene under salt-stress condition and red-light treatment. For the data normalization under drought-stress treatment, FaDBP is the recommended reference gene with the highest expression stability. FaHISTH4 was observed to be the best reference gene for data normalization under heat stress and blue-light treatment. This work provides information on selecting reference genes for accurate gene expression analyses of target genes in strawberry leaves under various abiotic stress and light-quality conditions.

Highlights

Abiotic environmental stresses can give rise to multiple morphological, biochemical, and molecular changes that negatively affect plant growth and productivity [1]
Seven reference genes that were previously reported by our group were used as candidate reference genes [10]
We systematically evaluated seven candidate reference genes based on their expression stability in leaf tissue subjected to different external conditions

Summary

Introduction

Abiotic environmental stresses can give rise to multiple morphological, biochemical, and molecular changes that negatively affect plant growth and productivity [1]. Various physiological mechanisms in plants participate in response to stress, including the regulation of biological membrane stability, the regulation of hormone synthesis and accumulation, and the activity of phenolic enzymes [2]. The expression changes of stress tolerance genes have contributed to improve the adaptation of plants to environmental conditions [4,5]. The identification of abiotic stress-related genes in plants can provide useful information about the molecular mechanisms underlying responses to abiotic stress. Recent research has proposed the roles of light quality in modulating tolerance to abiotic and biotic stresses by inducing the accumulation of plant pigments and regulating the activities of major ROS scavengers [6,7]. An integrated analysis of light quality and stress could provide more information about the cross-talk between the light environment and external stress conditions

Methods

Results

Discussion

Conclusion