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10.1084/jem.20171626 http://scihub22266oqcxt.onion/10.1084/jem.20171626 C5839763!5839763 !29436394     free     free     free Warning: file_get_contents(https://eutils.ncbi.nlm.nih.gov/entrez/eutils/elink.fcgi?dbfrom=pubmed&id=29436394 &cmd=llinks): Failed to open stream: HTTP request failed! 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Optimized RNP transfection for highly efficient CRISPR/Cas9-mediated gene knockout in primary T cells |pdf=|usr=}}{{29436394 }} gab.com Text Optimized RNP transfection for highly efficient CRISPR/Cas9-mediated gene knockout in primary T cells JO: J Exp Med http://www.kidney.de/pget.php?&tw=1&pmid=29436394 #FOAvip CRISPR (clustered, regularly interspaced, short palindromic repeats)/Cas9 (CRISPR-associated protein 9) has become the tool of choice for generating gene knockouts across a variety of species. The ability for efficient gene editing in primary T cells not only represents a valuable research tool to study gene function but also holds great promise for T cell-based immunotherapies, such as next-generation chimeric antigen receptor (CAR) T cells. Previous attempts to apply CRIPSR/Cas9 for gene editing in primary T cells have resulted in highly variable knockout efficiency and required T cell receptor (TCR) stimulation, thus largely precluding the study of genes involved in T cell activation or differentiation. Here, we describe an optimized approach for Cas9/RNP transfection of primary mouse and human T cells without TCR stimulation that results in near complete loss of target gene expression at the population level, mitigating the need for selection. We believe that this method will greatly extend the feasibly of target gene discovery and validation in primary T cells and simplify the gene editing process for next-generation immunotherapies. Twit Text FOAVip Optimized RNP transfection for highly efficient CRISPR/Cas9-mediated gene knockout in primary T cells ????J Exp Med http://www.kidney.de/pget.php?&tw=1&pmid=29436394 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=29436394 #FOAvip English Wikipedia <ref>{{Cite pmid|29436394 }}</ref> Optimized RNP transfection for highly efficient CRISPR/Cas9-mediated gene knockout in primary T cells #MMPMID29436394 Seki A ; Rutz S J Exp Med 2018[Mar]; 215 (3 ): 985-997 PMID29436394 show ga CRISPR (clustered, regularly interspaced, short palindromic repeats)/Cas9 (CRISPR-associated protein 9) has become the tool of choice for generating gene knockouts across a variety of species. The ability for efficient gene editing in primary T cells not only represents a valuable research tool to study gene function but also holds great promise for T cell-based immunotherapies, such as next-generation chimeric antigen receptor (CAR) T cells. Previous attempts to apply CRIPSR/Cas9 for gene editing in primary T cells have resulted in highly variable knockout efficiency and required T cell receptor (TCR) stimulation, thus largely precluding the study of genes involved in T cell activation or differentiation. Here, we describe an optimized approach for Cas9/RNP transfection of primary mouse and human T cells without TCR stimulation that results in near complete loss of target gene expression at the population level, mitigating the need for selection. We believe that this method will greatly extend the feasibly of target gene discovery and validation in primary T cells and simplify the gene editing process for next-generation immunotherapies. |*Gene Knockout Techniques [MESH] |*Transfection [MESH] |Animals [MESH] |CRISPR-Associated Protein 9/*metabolism [MESH] |CRISPR-Cas Systems/*genetics [MESH] |Cells, Cultured [MESH] |Electroporation [MESH] |Female [MESH] |HEK293 Cells [MESH] |Humans [MESH] |Lymphocyte Activation/immunology [MESH] |Mice, Inbred C57BL [MESH] |RNA, Guide, CRISPR-Cas Systems/metabolism [MESH] |Retroviridae/metabolism [MESH] |Ribonucleoproteins/*metabolism [MESH]Optimized RNP transfection for highly efficient CRISPR/Cas9-mediated g(epmc).pdf DeepDyve Pubget Overpricing