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A novel method to achieve high yield of total RNA from zebrafish for expression studies

Saurabh Shukla, Reena C. Jhamtani, M. S. Dahiya, Rakhi Agarwal*

Abstract


Gene expression studies require high-quality intact total RNA for real-time PCR analysis. Degraded or impure sample can provide disastrous results in the subsequence process. RNA is very unstable and can be contaminated easily by ribonuclease (RNase), therefore utmost quality control checks are required to ensure best quality RNA with high yield. Herein, we have come out with a protocol to isolate total RNA from liver, kidney and brain of adult zebrafish using a commercially available chemical denaturant and subsequent clean-up to remove traces of DNA and impurities. Kidney tissue gives maximum yield of total RNA concentration i.e. 135.8786 ng/µL. Purity of isolated RNA was observed with an absorbance ranging from 1.9 to 2.0 observed in all samples. The whole procedure was repeatedly performed to get satisfactory results. Throughout these procedures there are numerous quality control checks to ensure that the sample is neither degraded nor contaminated.


Keywords


Gene Expression; Ribozol; RT-PCR; Total RNA; Zebrafish

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References


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DOI: http://dx.doi.org/10.21746/ijbio.2017.05.004

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