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Maximizing CRISPRi efficacy and accessibility with dual-sgRNA libraries and optimal effectors.

Replogle JM, Bonnar JL, Pogson AN, Liem CR, Maier NK, Ding Y, Russell BJ, Wang X, Leng K, Guna A, Norman TM, Pak RA, Ramos DM, Ward ME, Gilbert LA, Kampmann M, Weissman JS, Jost M
eLife. 2022 Dec 28;11:e81856. doi: 10.7554/eLife.81856. (Link opens in a new window) PubMed (Link opens in a new window) Article

Plasmids from Article

ID Plasmid Purpose
187239pJR98CR3 constant region - hU6 sgRNA promoter flanked by BsmBI sites
187240pJR100Lentiviral sgRNA vector for Perturb-seq with mU6 sgRNA promoter, CR1 constant region with CS1 capture sequence in stem loop, and UCOE EF1alpha driving PURO-BFP marker expression
187241pJR101Lentiviral sgRNA vector for Perturb-seq with mU6 sgRNA promoter, CR1 constant region with CS1 capture sequence in stem loop, and UCOE EF1alpha driving PURO-GFP marker expression
187242pJR103Lentiviral sgRNA vector with mU6 sgRNA promoter, CR1 constant region, and UCOE EF1alpha driving PURO-BFP marker expression
187243pJR104Lentiviral sgRNA vector with mU6 sgRNA promoter, CR1 constant region, and UCOE EF1alpha driving PURO-GFP marker expression
187244pJR106Lentiviral sgRNA vector for CROP-seq with mU6 sgRNA promoter, CR1 constant region, and UCOE EF1alpha driving PURO-BFP marker expression
187245pJR107Lentiviral sgRNA vector for CROP-seq with mU6 sgRNA promoter, CR1 constant region, and UCOE EF1alpha driving PURO-GFP marker expression
187246hCRISPRi_dual_1_2Ultra-compact, human genome-wide CRISPRi library targeting each gene with a single element encoding a dual sgRNA cassette.
187247hCRISPRi_dual_3_4Ultra-compact, human genome-wide CRISPRi library targeting each gene with a single element encoding a dual sgRNA cassette.
187248hCRISPRi_dual_5_6Ultra-compact, human genome-wide CRISPRi library targeting each gene with a single element encoding a dual sgRNA cassette.
187249hCRISPRa_dual_1_2Ultra-compact, human genome-wide CRISPRi library targeting each gene with a single element encoding a dual sgRNA cassette.
187250hCRISPRa_dual_3_4Ultra-compact, human genome-wide CRISPRi library targeting each gene with a single element encoding a dual sgRNA cassette.
187251hCRISPRa_dual_5_6Ultra-compact, human genome-wide CRISPRi library targeting each gene with a single element encoding a dual sgRNA cassette.
188765pHR-UCOE-SFFV-dCas9-XTEN80-KRAB(Kox1)-P2A-EGFPdCas9 with a C-term HA-2xNLS-XTEN80(linker)-KRAB(Kox1)
188766pHR-UCOE-EF1a-Zim3-dCas9-P2A-mCherrydCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188767pHR-UCOE-SFFV-Zim3-dCas9-P2A-BFPdCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188768pHR-UCOE-SFFV-Zim3-dCas9-P2A-HygrodCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188769pHR_UCOE-EF1a-dCas9-XTEN80-KRAB(Kox1)-IRES-mCherrydCas9 with a C-term HA-2xNLS-XTEN80(linker)-KRAB(Kox1)
188770pHR_UCOE-SFFV-dCas9-XTEN80-KRAB(Kox1)-IRES-mCherrydCas9 with a C-term HA-2xNLS-XTEN80(linker)-KRAB(Kox1)
188771pHR-UCOE-EFS-dCas9-XTEN80-KRAB(Kox1)-P2A-EGFPdCas9 with a C-term HA-2xNLS-XTEN80(linker)-KRAB(Kox1)
188772pHR-UCOE-CMV-dCas9-XTEN80-KRAB(Kox1)-P2A-EGFPdCas9 with a C-term HA-2xNLS-XTEN80(linker)-KRAB(Kox1)
188773pHR-UCOE-EF1a-Zim3-dCas9-loxP-P2A-EGFP-loxPdCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188774pHR-UCOE-SFFV-Zim3-dCas9-loxP-P2A-EGFP-loxPdCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188775pHR-UCOE-EF1a-Zim3-dCas9-BFPdCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188776pHR-UCOE-SFFV-Zim3-dCas9-BFPdCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188777pHR-UCOE-EF1a-Zim3-dCas9-P2A-BFPdCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188778pHR-UCOE-EF1a-Zim3-dCas9-P2A-GFPdCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188779pHR-UCOE-SFFV-Zim3-dCas9-P2A-mCherrydCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS
188898pHR-UCOE-SFFV-dCas9-P2A-EGFPHuman lentiviral vector for expression of dCas9 with a C-terminal HA-2xNLS and GFP linked by a P2A site from a SFFV promoter with an upstream ubiquitous chromatin opening element
188899pHR-UCOE-SFFV-Zim3-dCas9-P2A-EGFPdCas9 with an N-term Zim3 KRAB-NLS fusion, a C-term HA-2xNLS, and GFP linked by a P2A site from a SFFV promoter with an upstream ubiquitous chromatin opening element
188900pHR-UCOE-SFFV-EGFPHuman lentiviral vector for expression of EGFP from a SFFV promoter with an upstream ubiquitous chromatin opening element
188901pHR-UCOE-SFFV-SID4x-dCas9-XTEN80-KRAB(Kox1)-P2A-EGFPdCas9 with an N-term Sid4x fusion, a C-term HA-2xNLS-XTEN80(linker)-KRAB(Kox1) fusion, and GFP linked by a P2A site from a SFFV promoter with an upstream ubiquitous chromatin opening element
188902pHR-UCOE-SFFV-dCas9-KRAB(Kox1)-MeCP2-P2A-EGFPdCas9 with a C-term NLS-KRAB(Kox1)-MeCP2 fusion (no linker), and GFP linked by a P2A site from a SFFV promoter with an upstream ubiquitous chromatin opening element

Antibodies from Article