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Versatile and efficient non-viral integration of large transgenes in human T cells via CRISPR knock-in and engineered integrases

Kassing I, Kath J, Nitulescu A-M, Glaser V, Hartmann LM, Pu Y, Huth L, Kārkliņš R, Shaji S, Ringel AR, Pouzolles M, Stein M, Ibrahim DM, Wagner DL
bioRxiv 2025.09.10.675267 (Link opens in a new window) Article

Plasmids from Article

ID Plasmid Purpose
247037PASTA-1_pUC19_HDR-Bxb1-attG_TRAC_for-Cas12aEncodes HDR-template for Cas12a-mediated Knock-In of Bxb1-attG (landing pad) into TRAC gene (exon1). Template region to be PCR-amplified.
247038PASTA-2_trunc-pUC19_INT-Bxb1-attV_P2A_CD19CAR_bGH-pAIntegration-Plasmid with Bxb1-attV (docking site) for bxb1-mediated integration of a CD19-CAR. Targeted whole plasmid insertion after maxiprep + AMPure bead purification.
247039PASTA-3_pUC19_HDR-Pa01-attG_TRAC_for-Cas12aEncodes HDR-template for Cas12a-mediated Knock-In of size-reduced Pa01-attG (landing pad) into TRAC gene (exon1). Template region to be PCR-amplified.
247040PASTA-4_pUC19_INT-Pa01-attV_P2A_CD19CAR_bGH-pAIntegration-Plasmid with Pa01-attV (docking site) for Pa01-mediated integration of a CD19-CAR. Targeted whole plasmid insertion after maxiprep + AMPure bead purification.
247041PASTA-5_pUC19_IVT-ePa01Plasmid for in vitro transcription (IVT) of enhanced Pa01 (ePa01) mRNA. Use indicated PCR-primers for correction of T7-mutation and addition of poly-A-tail.

Antibodies from Article