Generation of Genetic Knockouts in Myeloid Cell Lines Using a Lentiviral CRISPR/Cas9 System
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.
KeywordsCRISPR Lentivirus Myeloid THP-1 Immortalized BMDM CRISPR validation
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.