Reference Gene Selection for Quantitative Real-Time PCR in Chrysanthemum Subjected to Biotic and Abiotic Stress

• Published in: Molecular biotechnology Vol. 49; no. 2; pp. 192 - 197
• Main Authors: Gu, Chunsun, Chen, Sumei, Liu, Zhaolei, Shan, Hong, Luo, Huolin, Guan, Zhiyong, Chen, Fadi
• Format: Journal Article
• Language: English
• Published: New York Springer-Verlag 01.10.2011; Humana Press Inc; Springer; Springer Nature B.V
• Subjects: actin; Aphididae; Aphidoidea; Biochemistry; Biological and medical sciences; Biological Techniques; Biotechnology; Cell Biology; Chemistry; Chemistry and Materials Science; Chrysanthemum; Chrysanthemum - genetics; Chrysanthemum - metabolism; computer software; flooded conditions; Fundamental and applied biological sciences. Psychology; Gene expression; Gene Expression Profiling - standards; genes; Genes, Plant - genetics; Genomics; glyceraldehyde-3-phosphate dehydrogenase; heat; heat stress; Human Genetics; phosphoglycerate kinase; photosystem I; Plant biology; Plant Proteins - genetics; Plant Proteins - metabolism; Plastids; Polymerase chain reaction; Protein Science; protein subunits; quantitative polymerase chain reaction; Real-Time Polymerase Chain Reaction - methods; Reference Standards; Stress, Physiological - genetics; tubulin; ubiquitin; Quantitative real-time PCR; Normalization; Chrysanthemum; Reference genes; Selection; Standardization; Environmental factor; Compositae; Real time; Stress; Abiotic factor; Polymerase chain reaction; Gene; Dicotyledones; Angiospermae; Spermatophyta; Quantitative analysis
• ISSN: 1073-6085; 1559-0305
• DOI: 10.1007/s12033-011-9394-6

Quantitative real-time PCR (RT-qPCR) is a reliable method for assessing gene expression, provided that suitable reference genes are included to normalize the data. The stability of expression of eight potential reference genes, namely, tubulin (alpha-2,4 tubulin), actin, EF1α (elongation factor 1α), UBC (ubiquitin C), GAPDH (glyceraldehyde-3-phosphate dehydrogenase), psaA (photosynthesis-related plastid gene representing photosystem I), PP2Acs (catalytic subunit of protein phosphatase 2A), and PGK (phosphoglycerate kinase), was assessed in chrysanthemum plants subjected to aphid infestation, heat stress or waterlogging stress using geNorm software. The widely used reference gene EF1α performed well for aphid infested plants but poorly for waterlogged ones. The catalytic subunit of protein phosphatase 2A (PP2Acs) was the best performing one during heat and waterlogging stress, but was the worst during aphid infestation. The commonly used reference gene actin was generally the least stable of the set. No single gene was suitable for normalization on its own. The choice of reference gene(s) is an important factor in gene expression studies based on RT-qPCR.