Selection of appropriate reference genes for RT-qPCR analysis under abiotic stress and hormone treatment in celery

Kai Feng; Jie-Xia Liu; Guo-Ming Xing; Sheng Sun; Sen Li; Ao-Qi Duan; Feng Wang; Meng-Yao Li; Zhi-Sheng Xu; Ai-Sheng Xiong

doi:10.7717/peerj.7925

Selection of appropriate reference genes for RT-qPCR analysis under abiotic stress and hormone treatment in celery

PeerJ. 2019 Oct 24:7:e7925. doi: 10.7717/peerj.7925. eCollection 2019.

Authors

Affiliations

¹ State Key Laboratory of Crop Genetics and Germplasm Enhancement, Ministry of Agriculture and Rural Affairs Key Laboratory of Biology and Germplasm Enhancement of Horticultural Crops in East China, College of Horticulture, Nanjing Agricultural University, Nanjing, China.
² Collaborative Innovation Center for Improving Quality and Increase Profits of Protected Vegetables in Shanxi, Taigu, China.

Abstract

Celery is one of the most important vegetable crop and its yield and quality is influenced by many environmental factors. Researches on gene expression not only help to unravel the molecular regulatory mechanism but also identify the key genes in the biological response. RT-qPCR is a commonly used technology to quantify the gene expression. Selecting an appropriate reference gene is an effective approach to improve the accuracy of RT-qPCR assay. To our knowledge, the evaluation of reference genes under different treatments in celery has not been reported yet. In this study, the expression stabilities of eight candidate reference genes (ACTIN, eIF-4α , GAPDH, TBP, TUB-A, UBC, TUB-B, and EF-1α ) under abiotic stresses (heat, cold, drought, and salt) and hormone treatments (SA, MeJA, GA, and ABA) were detected. The expression stabilities of candidate genes were compared and ranked by geNorm, NormFinder, BestKeeper, ΔCt, and RefFinder programs. The results calculated by different programs were not completely consistent. Considering the comprehensive analysis results, ACTIN was the most stable reference gene and TUB-B showed the worst expression stabilities under the selected abiotic stress and hormone treatments in celery. The reliability of reference genes was further confirmed by the normalization of CAT1 gene under drought stress. This study presented evidences and basis to select the appropriate reference genes under different treatments in celery.

Keywords: Abiotic stress; Celery; Expression stability; Hormone stimuli; RT-qPCR; Reference gene.

Grants and funding

This study was supported by the Jiangsu Agricultural Science and Technology Innovation Fund [CX(2018)2007], New Century Excellent Talents in University (NCET-11-0670); the National Natural Science Foundation of China (31272175); and the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD). There was no additional external funding received for this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.