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Selection and validation of reference genes by RT-qPCR under photoperiodic induction of flowering in sugarcane ( Saccharum spp.)
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da Silva Santos, Paulo H., Manechini, João R. Vieira, Brito, Michael S., Romanel, Elisson, Vicentini, Renato, Scarpari, Maximiliano, Jackson, Stephen D. and Pinto, Luciana R. (2021) Selection and validation of reference genes by RT-qPCR under photoperiodic induction of flowering in sugarcane ( Saccharum spp.). Scientific Reports, 11 (1). 4589. doi:10.1038/s41598-021-83918-2 ISSN 2045-2322.
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WRAP-Selection-validation-genes-photoperiodic-induction-sugarcane-2021.pdf - Published Version - Requires a PDF viewer. Available under License Creative Commons Attribution 4.0. Download (1285Kb) | Preview |
Official URL: https://doi.org/10.1038/s41598-021-83918-2
Abstract
Although reference genes have previously been used in the expression analysis of genes involved in sugarcane flowering they had not been experimentally validated for stability and consistency of expression between different samples over a wide range of experimental conditions. Here we report the analysis of candidate reference genes in different tissue types, at different temporal time-points, in both short and long day photoperiodic treatments. The stability of the candidate reference genes in all conditions was evaluated with NormFinder, BestKeeper, and RefFinder algorithms that complement each other for a more robust analysis. As the Normfinder algorithm was more appropriate for our experimental conditions, greater emphasis was placed on Normfinder when choosing the most stable genes. UBQ1 and TUB were shown to be the most stable reference genes to use for normalizing RT-qPCR gene expression data during floral induction, whilst 25SrRNA1 and GAPDH were the least stable. Their use as a reference gene pair was validated by analyzing the expression of two differentially expressed target genes (PIL5 and LHP1). The UBQ1/TUB reference genes combination was able to reveal small significant differences in gene expression of the two target genes that were not detectable when using the least stable reference gene combination. These results can be used to inform the choice of reference genes to use in the study of the sugarcane floral induction pathway. Our work also demonstrates that both PIL5 and LHP1 are significantly up-regulated in the initial stages of photoperiodic induction of flowering in sugarcane.
Item Type: | Journal Article | |||||||||
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Subjects: | Q Science > QK Botany S Agriculture > SB Plant culture |
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Divisions: | Faculty of Science, Engineering and Medicine > Science > Life Sciences (2010- ) | |||||||||
SWORD Depositor: | Library Publications Router | |||||||||
Library of Congress Subject Headings (LCSH): | Sugarcane, Photoperiodism | |||||||||
Journal or Publication Title: | Scientific Reports | |||||||||
Publisher: | Nature Publishing Group UK | |||||||||
ISSN: | 2045-2322 | |||||||||
Official Date: | 25 February 2021 | |||||||||
Dates: |
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Volume: | 11 | |||||||||
Number: | 1 | |||||||||
Article Number: | 4589 | |||||||||
DOI: | 10.1038/s41598-021-83918-2 | |||||||||
Status: | Peer Reviewed | |||||||||
Publication Status: | Published | |||||||||
Reuse Statement (publisher, data, author rights): | ** From Springer Nature via Jisc Publications Router ** History: received 27-09-2020; accepted 03-02-2021; registration 10-02-2021; online 25-02-2021; pub-electronic 25-02-2021; collection 12-2021. ** Licence for this article: http://creativecommons.org/licenses/by/4.0/ | |||||||||
Access rights to Published version: | Open Access (Creative Commons) | |||||||||
Date of first compliant deposit: | 5 March 2021 | |||||||||
Date of first compliant Open Access: | 9 March 2021 | |||||||||
RIOXX Funder/Project Grant: |
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