Identification of reliable reference genes for quantitative real-time PCR normalization in pitaya
Canbin Chen,
Jingyu Wu,
Qingzhu Hua,
Noemi Tel-Zur,
Fangfang Xie,
Zhike Zhang,
Jianye Chen,
Rong Zhang,
Guibing Hu,
Jietang Zhao,
Yonghua Qin
Affiliations
Canbin Chen
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Jingyu Wu
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Qingzhu Hua
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Noemi Tel-Zur
French Associates Institute for Agriculture and Biotechnology of Drylands, The J. Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev
Fangfang Xie
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Zhike Zhang
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Jianye Chen
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Rong Zhang
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Guibing Hu
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Jietang Zhao
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Yonghua Qin
State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources/Guangdong Provincial Key Laboratory of Postharvest Science of Fruits and Vegetables/Key Laboratory of South China Horticultural Crop Biology and Germplasm Enhancement, Ministry of Agriculture, College of Horticulture, South China Agricultural University
Abstract Background A suitable reference gene is an important prerequisite for guarantying accurate and reliable results in quantitative real-time PCR (qRT-PCR) analyses. However, there is no absolute universality in reference genes among different species. It’s hard to find an ideal reference gene to fit for different tissues and growth periods. Pitaya (Hylocereus) is commercially produced as a new fruit crop at a large scale in tropical and subtropical regions. To date, there is no report on the identification of the most reliable reference genes for qRT-PCR normalization in pitaya. Results In this study, six candidate reference genes i.e. Actin(1), GAPDH, UBC(1), UBC(2) EF1-α(1) and histone(1) were selected from thirty-nine typical candidate reference genes to determine the most stable reference genes for qRT-PCR normalization in different tissues, temperature stresses and fruit developmental stages of pitaya. Among the six candidate reference genes, Actin(1) and EF1-α(1) were the most stable gene according to calculations of three statistical methods (GeNorm, NormFinder and BestKeeper) while UBC(1) and UBC(2) showed the lowest expression stability. The six candidate reference genes were further validated by comparing expression profiles of key genes related to betalain biosynthesis at flesh coloration stages of Guanhuahong (Hylocereus monacanthus) and Guanhuabai (H. undatus) pitayas. Actin(1) was recommended the best reference gene for accurate normalization of qRT-PCR data. Conclusions In this study, the stability of the selected reference genes for normalizing the qRT-PCR data were identified from pitaya. Actin(1) was the most stably expressed genes in different tissues and fruit developmental stages in pitaya. The present work provides the first data of reference gene identification for pitaya and will facilitate further studies in molecular biology and gene function on Hylocereus and other closely related species.
