Molecular Biology Reports

, Volume 39, Issue 2, pp 1831–1838

Selection of reference genes for quantitative real-time RT-PCR analysis in citrus

  • Jiawen Yan
  • Feirong Yuan
  • Guiyou Long
  • Lei Qin
  • Ziniu Deng
Article

DOI: 10.1007/s11033-011-0925-9

Cite this article as:
Yan, J., Yuan, F., Long, G. et al. Mol Biol Rep (2012) 39: 1831. doi:10.1007/s11033-011-0925-9

Abstract

Quantitative real-time reverse transcription polymerase chain reaction (qPCR) has become the preferred method for studying low-abundant mRNA expression. Appropriate application of qPCR in such studies requires the use of reference gene(s) as an internal control in order to normalize the mRNA levels between different samples for an exact comparison of gene expression levels. Expression of the reference gene should be independent from development stage, cell/tissue types, treatments and environmental conditions. Recognizing the importance of reference gene(s) in normalization of qPCR data, various reference genes have been evaluated for stable expression under specific conditions in various organisms. In plants, only a few of them have been investigated, and very few reports about such reference genes in citrus. In the present study, seven candidate reference genes (18SrRNA, ACTB, rpII, UBQI, UBQ10, GAPDH and TUB) were tested, and three of them (18SrRNA, ACTB and rpII) proved to be the most stable ones among six leaf samples of different citrus genotypes. The three candidate reference genes were further analyzed for their stability of expression in five different tissues, and the results indicated that they were not completely stable. It is commonly accepted that gene expression studies should be normalized using more than one reference gene. Based on our results, we propose the use of the mean result rendered by18SrRNA, ACTB and rpII as reference genes to normalize mRNA levels in qPCR analysis of diverse cultivars and tissues of citrus. These results may provide a guideline for future works on gene expression in citrus by using qPCR.

Keywords

CitrusReference genesqPCRGene expression

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Jiawen Yan
    • 1
    • 2
    • 3
  • Feirong Yuan
    • 1
    • 2
    • 3
  • Guiyou Long
    • 1
    • 2
  • Lei Qin
    • 1
  • Ziniu Deng
    • 1
    • 2
    • 3
  1. 1.Horticulture and Landscape CollegeHunan Agricultural UniversityFurong District, ChangshaChina
  2. 2.National Center for Citrus ImprovementChangshaChina
  3. 3.Hunan Provincial Key Laboratory of Crop Germplasm Innovation and UtilizationChangshaChina