Evaluation of housekeeping genes as references for quantitative real-time PCR analysis of gene expression in the murrel Channa striatus under high-temperature stress
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Quantitative real-time polymerase chain reaction is the most advanced method of quantifying gene expression studies; however, the significance of the obtained results strongly depends on the normalization of the data to compensate for differences between the samples. In the present study, expression analysis of six different constitutively expressed genes viz. 18S ribosomal RNA, glyceraldehyde-3-phosphate dehydrogenase (gapdh), beta actin (βactin), ribosomal binding protein L13, tubulin and TATA-box-binding protein (tbp) were carried out to test their efficacy as reference genes in three different tissues, namely liver, gill and muscle of murrel Channa striatus exposed to high temperature for variable time periods. The stability and suitability of the genes were determined by using bioinformatic tools: GeNorm, NormFinder and BestKeeper. Based on the results, tub/βactin could be used as the reference genes for liver and gill tissues and βactin/gapdh could be the reference genes for muscle tissues in Channa striatus under both short- and long-term thermal stress.
KeywordsReference genes Quantitative real-time PCR Normalization Channa striatus Thermal stress
This work was funded by the Indian Council of Agricultural Research under the National Fund for Basic, Strategic and Frontier Application Research in Agriculture (NFBSFARA) (currently renamed National Agricultural Science Fund, (NASF) (Project #AS: 2001) to BPM, SM. GKP and AM are Senior Research Fellows. The authors are thankful to the Director, KIIT School of Biotechnology, Bhubaneswar, and Director, ICAR-Central Inland Fisheries Research Institute, Barrackpore, for the facilities and encouragement.
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Conflict of interest
The authors declare that there is no conflict of interest.
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