Abstract
RNA interference (RNAi) is a mechanism regulating gene transcript levels either by transcriptional gene silencing or by posttranscriptional gene silencing, which act in the genome maintenance and the regulation of gene expression which is typically inferred from measuring transcript abundance. Nuclear “run-on” (or “run-off”) transcription assays have been used to obtain quantitative information about the relative rates of transcription of different genes in nuclei isolated from a particular tissue or organ. Basically, these assays exploit the activity of RNA polymerases to synthesize radiolabeled transcripts that then can be hybridized to filter-bound, cold, excess single-stranded DNA probes representing genes of interest. The protocol presented here streamlines, adapts, and optimizes nuclear run-on transcription assays for use in RNAi studies.
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Acknowledgments
The author would like to thank Dr. Enas Qudeimat and Dr. Mieke Van Lijsebettens for helpful comments on the manuscript.
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Khraiwesh, B. (2011). Using Nuclear Run-On Transcription Assays in RNAi Studies. In: Kodama, H., Komamine, A. (eds) RNAi and Plant Gene Function Analysis. Methods in Molecular Biology, vol 744. Humana Press. https://doi.org/10.1007/978-1-61779-123-9_14
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DOI: https://doi.org/10.1007/978-1-61779-123-9_14
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