Description:

CRISPR interference (CRISPRi) emerges as a potent tool, allowing targeted silencing of gene transcription in both prokaryotic and eukaryotic cells. The synergy of inactive Cas9 (dCas9) and the Krüppel-associated box (KRAB) repressor enables accurate and reversible gene silencing. Furthermore, the strategic utilization of the Human AAVS1 "safe harbor" site, located within intron 1 of the PPP1R12C gene, minimizes its impact on cellular functions.Our innovative pAAVS1-SA-T2A-Neo-CAG-dCas9-KRAB-pA donor vector seamlessly combines the strengths of CRISPRi and the AAVS1 safe harbor. Featuring a CAG promoter-controlled CRISPRi-KRAB fusion gene, this design ensures consistent and robust expression of the dCAS9/KRAB protein without disrupting cellular functions. The vector also strategically integrates the Neomycin selection marker with a splice acceptor (SA) site within the AAVS1 safe harbor locus, closely tying Neomycin expression to intron integration. This significantly reduces the risk of unintended off-target integrations during Puromycin selection. When employed with the pSpCAS9 nuclease & AAVS1 gRNA vector, the pAAVS1-SA-T2A-Neo-CAG-dCas9-KRAB-pA donor vector facilitates the seamless integration of a dCAS9/KRAB fusion protein into the AAVS1 safe harbor site. This approach, combined with specific gene-targeted gRNAs, enables precise and highly efficient repression of the target gene expression, offering an easy and consistent method for specific gene silencing.

Vector Information

This AAVS1 donor vector includes essential components to serve as a reliable integration marker in the AAVS1 Safe Harbor site. It incorporates a universal SA-T2A-NeoR cassette, facilitating the integration of a dCas9/KRAB fusion gene driven by a CAG promoter into the AAVS1 safe harbor site through antibiotic selection. This vector is suitable for generating CRISPRi stable cell lines.

IMPORTANT NOTICE

Store the vial at -20°C immediately upon receipt.

Specifications:

Product Name	pAAVS1-SA-T2A-Neo-CAG-dCas9-KRAB-pA Donor Vector
Shipping Condition	Room Temperature - 2 Day Shipping
Storage and Stability	Store at -20°C immediately upon receipt. This product is stable for 6 months when stored as directed.
Quality Control	This plasmid is sequence verified.
Restricted Use	For Research Use Only. Not for use in diagnostic or therapeutic procedures.