Enhancing gRNA Transcript levels by Reducing the Scaffold Poly-T Tract for Optimal SpCas9- and SaCas9-mediated Gene Editing

Chey YCJ, Gierus L, Lushington C, Arudkumar JC, Geiger A, Staker LG, Robertson LJ, Pfitzner C, Kennedy JG, Lee RHB, Godahewa GI, Thomas PQ, Adikusuma F
bioRxiv 2024.07.19.604224 Article

Plasmids from Article

ID	Plasmid	Purpose
178799	PX459v3	SpCas9 with 2A-Puro, and a golden gate cloning backbone for sgRNA. sgRNA scaffold seqeuence has been modified for increased sgRNA expression (v3).
178800	PX459v3-eSpCas9(1.1)	High-fidelity eSpCas9(1.1) with 2A-Puro, and a golden gate cloning backbone for sgRNA. sgRNA scaffold seqeuence has been modified for increased sgRNA expression (v3).
178801	PX459v3-SpCas9-HF1	High-fidelity SpCas9-HF1 with 2A-Puro, and a golden gate cloning backbone for sgRNA. sgRNA scaffold seqeuence has been modified for increased sgRNA expression (v3).
178802	SaCas9v2-Puro	SaCas9 with 2A-Puro, and a cloning backbone for sgRNA.
178813	SaCas9v3-Puro	SaCas9 with 2A-Puro, and a cloning backbone for sgRNA. sgRNA scaffold seqeuence has been modified for increased sgRNA expression (v3).
226955	ABEmax-Puro V2	ABEmax plasmid enabling A:T to G:C base editing using a single plasmid.
226957	PX458 V3	CBh-SpCas9-2A-GFP, and hU6-sgRNA (Sp) with a BbsI golden gate cloning backbone for sgRNA. sgRNA uses 3TC scaffold for increased sgRNA expression.
226958	PDG459 V3	CBh-SpCas9-2A-Puro, and 2X hU6-sgRNA (Sp) with BbsI golden gate cloning backbone dual gRNAs. sgRNA uses 3TC scaffold for increased sgRNA expression.
226959	PDG458 V3	CBh-SpCas9-2A-GFP, and 2X hU6-sgRNA (Sp) with BbsI golden gate cloning backbone dual gRNAs. sgRNA uses 3TC scaffold for increased sgRNA expression.
226960	DG SaCas9 puro V3	CBh-SaCas9-2A-Puro, and 2X hU6-sgRNA (Sa) with BbsI golden gate cloning backbone for dual gRNAs. sgRNA uses 3TC scaffold for increased sgRNA expression.
226961	DG SaCas9 GFP V3	CBh-SaCas9-2A-GFP, and 2X hU6-sgRNA (Sa) with BbsI golden gate cloning backbone for dual gRNAs. sgRNA uses 3TC scaffold for increased sgRNA expression.
226962	SaCas9 GFP V3	CBh-SaCas9-2A-GFP, and hU6-sgRNA (Sa) with a BbsI golden gate cloning backbone for sgRNA. sgRNA uses 3TC scaffold for increased sgRNA expression.
226963	PX458 eSp(1.1) V3	CBh-eSpCas9(1.1)-2A-GFP, and hU6-sgRNA (Sp) with a BbsI golden gate cloning backbone for sgRNA. sgRNA uses 3TC scaffold for increased sgRNA expression.
226964	PDG458 eSp(1.1) V3	CBh-eSpCas9(1.1)-2A-GFP, and 2X hU6-sgRNA (Sp) with BbsI golden gate cloning backbone dual gRNAs. sgRNA uses 3TC scaffold for increased sgRNA expression.
226965	Pdg459-eSp(1.1) V3	CBh-eSpCas9(1.1)-2A-Puro, and 2X hU6-sgRNA (Sp) with a BbsI golden gate cloning backbone for sgRNA. sgRNA uses 3TC scaffold for increased sgRNA expression.