DCAS9P300VP

Sigma CRISPR dCAS9-p300V Activator Lenti Plasmid

• UNSPSC Code: 12352200
• NACRES: NE.02

Properties

• recombinant: expressed in E. coli
• Quality Level: 200
• form: liquid
• packaging: vial of 50 μL
• concentration: 20 ng/μL in TE buffer; DNA (1μg of plasmid DNA)
• application(s): CRISPR
• shipped in: dry ice
• storage temp.: −20°C

Description

General description

This gene activation system is based on a fusion of inactive Cas9 (dCas9) to the catalytic histone acetyltransferase (HAT) core domain of the human E1A-associated protein p300. The dCas9-p300 activator lenti plasmids use the EF1 alpha promoter for strong expression of dCas9-P300 and blasticidin linked by a 2A peptide (EF1a-dCas9-P300-2A-Blasticidin) allowing for easy selection following successful transfection or transduction. Use Sigma′s lentiviral dCas9-P300 lenti plasmid for generation of lentiviral particles and efficient production of stable cell lines expressing dCas9-P300 for CRISPR based gene activation. The dCas9-P300 lenti plasmid is one part of a two part CRISPR system with individual dCas9-P300 and gRNA expression vectors.

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Application

• Functional Genomics/Target Validation
• Epigenetic Modification
• Transcriptional Activation
• Manufacture of dCas9-P300 expressing lentiviral particles

Features and Benefits

The Sigma CRISPR dCas9p300V plasmid co-expresses p300-HAT and Blasticidin, allowing for blasticidin based selection of cells expressing dCas9p300. gRNAs can successfully direct nuclease-deficient Cas9 (dCas9) fused to p300 HAT catalytic domain to increase levels of histone acetylation and endogenous gene expression. The dCas9-p300 histone acetylation approach represents a distinct mechanism of action relative to dCas9-VP64 or other similar gene activation motifs.

Principle

CRISPR/Cas systems are employed by bacteria and archaea as a defense against invading viruses and plasmids. Recently, the type II CRISPR/Cas system from the bacterium Streptococcus pyogenes has been engineered to function in eukaryotic systems using two molecular components: a single Cas9 protein and a non-coding guide RNA (gRNA). Mutations to the catalytic domains, RuvC and HnH, render it inactive as a nuclease yet still allow for the protein to be programmed to target specific sequences of DNA with a single gRNA. A fusion of dCas9 to the catalytic histone acetyltransferase (HAT) core domain of the human E1A-associated protein p300 induces transcription by releasing DNA from its heterochromatin state allowing for continued and robust gene expression by endogenous cellular machinery. The dCas9-p300 CRISPR Gene Activator represents a distinct mechanism of action relative to dCas9-VP64 or other similar gene activation motifs

Legal Information

CRISPR Use License Agreement

Lentiviral, WPRE, and Evrogen Fluorophore License

Safety Information

• Storage Class Code: 10 - Combustible liquids
• WGK: WGK 2
• Flash Point(F): Not applicable
• Flash Point(C): Not applicable