Differential Expression Analysis of Reference Genes in Pineapple (Ananas comosus L.) during Reproductive Development and Response to Abiotic Stress, Hormonal Stimuli
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Pineapple (Ananas comosus L.), a popular tropical fruit, is a good model for evolutionary analysis and genetic research on adaptation to drought habitats, multiple fruits, and crassulacean acid metabolism (CAM) photosynthesis. Reliable reference genes for the normalization of the levels of development-related and/or stress-responsive genes is important for elucidating the mechanisms of plant development as well as their adaptation to various environments. In this study, ten candidate reference genes were selected, and the expression stability of each gene was assessed across 105 pineapple samples, consisting of different reproductive organs (stigma, petal, sepal, ovule, anther, flower, and fruitlet), abiotic stress (salinity, drought, and cold), and hormones (abscisic acid, ethylene, jasmonic acid, and salicylic acid). Our results revealed that PP2A and UBQ were stably expressed during pineapple reproductive development. PP2A and CYC were the stable reference genes across abiotic stress, whereas RAN and EF1α/PP2A was the best candidates for various hormones. To validate the feasibility of using these stably expressed genes for further experiments, we evaluated the expression profile of AcMYB30 across different samples. Our results provide important insights on the growth and development of the pineapple plant as well as information on stress-tolerance genes and stress-signaling pathways in this important fruit.
KeywordsPineapple (Ananas comosus L.) Reference gene Reproductive development Abiotic stress Hormonal stimulus Gene expression
Crassulacean acid metabolism
Quantitative real-time PCR
This project was funded by the China Postdoctoral Science Foundation (2018 M632564), the National Natural Science Foundation of China (31800262), the Special Fund for Science and Technology Innovation in FAFU (KFA17439A), the Weng Hongwu Academic Innovation Research Fund of Peking University, the Natural Science Foundation of Fujian Province (2017 J01601), and the National Natural Science Foundation of China (U1605212).
X.N. and Y.Q. initiated and designed the research. X.N., B.H., H.C., X.H., and Z.S. performed the experiments. X.N., H.C., L.Z., and D.S. analyzed the data. X.N., H.C., and S.V.G.N. Priyadarshani contributed reagents/materials/analytic tools. X.N. wrote the paper. X.N. and Y.Q. revised the paper.
Compliance with Ethical Standards
Conflict of Interests
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
- Bustin SA (2002) Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems. J Mol Endocrinol 29Google Scholar
- Clement CR, de Cristo-Araújo M, d’Eeckenbrugge GC, Alves Pereira A, Picanço-Rodrigues D (2010) Origin and domestication of native amazonian crops. Divers 2:72–106Google Scholar
- Gutierrez L, Mauriat M, Guenin S, Pelloux J, Lefebvre JF, Louvet R, Rusterucci C, Moritz T, Guerineau F, Bellini C, Van Wuytswinkel O (2008a) The lack of a systematic validation of reference genes: a serious pitfall undervalued in reverse transcription-polymerase chain reaction (RT-PCR) analysis in plants. Plant Biotechnol J 6:609–618CrossRefGoogle Scholar
- Ma J, HY WC, Liu H, Hu Z (2012) Cloning and selection of reference genes for gene expression studies in Ananas comosus. African J Biotech 11:7424–7433Google Scholar
- Martins MQ, Fortunato AS, Rodrigues WP, Partelli FL, Campostrini E, Lidon FC, DaMatta FM, Ramalho JC, Ribeiro-Barros AI (2017) Selection and validation of reference genes for accurate RT-qPCR data normalization in Coffea spp. under a climate changes context of interacting elevated [CO2] and temperature. Front Plant Sci 8:307Google Scholar
- Michniewicz M, Zago MK, Abas L, Weijers D, Schweighofer A, Meskiene I, Heisler MG, Ohno C, Zhang J, Huang F, Schwab R, Weigel D, Meyerowitz EM, Luschnig C, Offringa R, Friml J (2007) Antagonistic regulation of PIN phosphorylation by PP2A and PINOID directs auxin flux. Cell 130:1044–1056CrossRefGoogle Scholar
- Smith MK, Ko HL, Hamill SD, Sanewski GM, Graham MW, Bartholomew DP, Paull RE, Rohrbach KG (2003) The pineapple: botany, production and uses. 1–320Google Scholar
- Wang C, Cui HM, Huang TH, Liu TK, Hou XL, Li Y (2016) Identification and validation of reference genes for RT-qPCR analysis in non-heading Chinese cabbage flowers. Front Plant Sci 7:811Google Scholar