Abstract
The application of CRISPR (clustered regularly interspaced short palindromic repeats)-Cas9 technology with pooled guide RNA libraries enables genome-wide screening, which has some advantages over other screening methods using chemical DNA mutagens for inducing genetic changes, RNA interference, or arrayed screens. Here we describe the use of genome-wide knockout and transcriptional activation screening enabling the CRISPR-Cas9 system to discover resistance mechanisms to CDK4/6 inhibition in bladder cancer along with next-generation sequencing (NGS) analysis. We will describe the approach for transcriptional activation in the bladder cancer cell line T24 and provide guidance on critical points during the experimental workflow.
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Acknowledgments
We thank Anuja Sathe and Zhichao Tong for their contribution to the preparation of this protocol.
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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
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Mantwill, K., Nawroth, R. (2023). Genome-Wide CRISPR Screening for the Identification of Therapy Resistance-Associated Genes in Urothelial Carcinoma. In: Hoffmann, M.J., Gaisa, N.T., Nawroth, R., Ecke, T.H. (eds) Urothelial Carcinoma. Methods in Molecular Biology, vol 2684. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3291-8_9
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DOI: https://doi.org/10.1007/978-1-0716-3291-8_9
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