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The Scientific World Journal
Volume 2014, Article ID 357052, 12 pages
Research Article

Validation of Novel Reference Genes for Reverse Transcription Quantitative Real-Time PCR in Drought-Stressed Sugarcane

1Federal University of Pernambuco (UFPE/CCB/Genética), 50670-420 Recife, PE, Brazil
2ABC Medical School, 09060-650 São Paulo, SP, Brazil
3Sugarcane Technology Center (CTC), 13400-970 Piracicaba, SP, Brazil
4Institute of Molecular Biosciences, Goethe University Frankfurt am Main, 60438 Frankfurt am Main, Germany

Received 14 January 2014; Revised 20 April 2014; Accepted 21 April 2014; Published 2 June 2014

Academic Editor: Juan J. Loor

Copyright © 2014 Roberta Lane de Oliveira Silva et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


One of the most challenging aspects of RT-qPCR data analysis is the identification of reliable reference genes. Ideally, they should be neither induced nor repressed under different experimental conditions. To date, few reference genes have been adequately studied for sugarcane (Saccharum spp.) using statistical approaches. In this work, six candidate genes (αTUB, GAPDH, H1, SAMDC, UBQ, and 25S rRNA) were tested for gene expression normalization of sugarcane root tissues from drought-tolerant and -sensitive accessions after continuous dehydration (24 h). By undergoing different approaches (GeNorm, NormFinder, and BestKeeper), it was shown that most of them could be used in combinations for normalization purposes, with the exception of SAMDC. Nevertheless three of them (H1, αTUB, and GAPDH) were considered the most reliable reference genes. Their suitability as reference genes validated the expression profiles of two targets (AS and PFPα1), related to SuperSAGE unitags, in agreement with results revealed by previous in silico analysis. The other two sugarcane unitags (ACC oxidase and PIP1-1), after salt stress (100 mM NaCl), presented their expressions validated in the same way. In conclusion, these reference genes will be useful for dissecting gene expression in sugarcane roots under abiotic stress, especially in transcriptomic studies using SuperSAGE or RNAseq approaches.