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Protocol for the saturation and multiplexing of genetic variants using CRISPR-Cas9

  1. Author:
    Sahu,Sounak
    Sullivan,Teresa
    Southon,Eileen
    Caylor,Dylan
    Geh, Josephine
    Sharan,Shyam

  2. Author Address

    Mouse Cancer Genetics Program, Centre for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA. Electronic address: sounak.sahu@nih.gov., Mouse Cancer Genetics Program, Centre for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA. Electronic address: sharans@mail.nih.gov.,
    1. Year: 2023
    2. Date: Nov 08
    3. Epub Date: 2023 11 08
  2. Journal: STAR Protocols
    1. 4
    2. 4
    3. Pages: 102702
  4. Type of Article: Article
  5. Article Number: 102702
  1. Abstract:

    Here, we present a multiplexed assay for variant effect protocol to assess the functional impact of all possible genetic variations within a particular genomic region. We describe steps for saturation genome editing by designing and cloning of single-guide RNA (sgRNA). We then detail steps for nucleofection of sgRNA, testing drug response on variants, and amplification of genomic DNA for next-generation sequencing. For complete details on the use and execution of this protocol, please refer to Sahu et al.1. Copyright © 2023. Published by Elsevier Inc.

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External Sources

  1. View Full Text
  2. DOI: 10.1016/j.xpro.2023.102702
  3. PMID: 37948185
  4. PII : S2666-1667(23)00669-X

Library Notes

  1. Fiscal Year: FY2023-2024

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