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Generation of Genetic Knockouts in Myeloid Cell Lines Using a Lentiviral CRISPR/Cas9 System

  • Paul J. BakerEmail author
  • Seth L. MastersEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1714)

Abstract

CRISPR/Cas9-based gene targeting allows deletion of a gene of interest from cultured cell lines. Due to difficulty in transiently transfecting hematopoetic cells with components required for this process, we have adopted a lentiviral system for delivery of the CRISPR/Cas9 components into myeloid cell lines. Here, we detail the process of knocking out genes from pools of cultured myeloid cells using this CRISPR/Cas9 system and describe methods of validating these knockout pools.

Keywords

CRISPR Lentivirus Myeloid THP-1 Immortalized BMDM CRISPR validation 

Notes

Acknowledgements

This work was supported by: Australian National Health and Medical Research Council (NHMRC) Project Grants (1057815 and 1099262) and Fellowship (S.L.M.); The Ian Potter Centre for Genomics and Personalized Medicine; and a Victorian State Government Operational Infrastructure Support Grant.

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Copyright information

© Springer Science+Business Media LLC 2018

Authors and Affiliations

  1. 1.Inflammation divisionThe Walter and Eliza Hall Institute of Medical ResearchParkvilleAustralia
  2. 2.Department of Medical BiologyThe University of MelbourneParkvilleAustralia

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