CRISPR08

CRISPR UNIVERSAL NEGATIVE CONTROL 3

• NACRES: NA.51
• UNSPSC Code: 41106609
• Concentration: 20 ng/μL in TE buffer; DNA (1μg of plasmid DNA)

Eigenschaften

• form: liquid
• Quality Segment: 200
• 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

Beschreibung

General description

Universal negative control CRISPRs have been designed not to recognize any sequence in the human, mouse or rat genome. A single vector format is provided which includes the Cas9 expression cassette and gRNA sequence. This vector includes GFP which is co-expressed from the same mRNA as the Cas9 protein via a 2A peptide linkage and enables for tracking of transfection efficiency or enrichment in cell populations via FACS.

Application

Functional Genomics/Target Validation

• Creation of gene knockouts in cell lines
• Creation of knock-in cell lines with promoters, fusion tags or reporters integrated into endogenous genes

Biochem/physiol Actions

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). The Cas9 endonuclease can be programmed with a single gRNA, directing a DNA double-strand break (DSB) at a desired genomic location. Similar to DSBs induced by zinc finger nucleases (ZFNs), the cell then activates endogenous DNA repair processes, either non-homologous end joining (NHEJ) or homology-directed repair (HDR), to heal the targeted DSB.

Features and Benefits

Allows for expression of Cas9 and GFP without specific targeting of the CRISPR/Cas9 system.

Physical form

Sigma U6-gRNA/CMV-Cas9-GFP plasmid expressing a non-targeting guide sequence supplied at a concentration of 20ng/ul in 50ul.

Preparation Note

Sigma CRISPR plasmid products are delivered as mini-prep aliquots, which may not be suitable for transfection into particular cell types. For best results, we advise maxi-prepping plasmids using endotoxin-free DNA purification kits prior to transfection.

Other Notes

1 vial containing 1ug of U6-gRNA/CMV-Cas9-GFP plasmid expressing a non-targeting guide sequence.

Typical transfection concentrations used in literature are in the ranges of 2.0 to 8.0ug of the single vector expressing the guideRNA and Cas9 . (All dosages above assume 0.5 to 1 million cells nucleofected)

Legal Information

CRISPR Use License Agreement

Lentiviral and WPRE License Agreements

Sicherheitshinweise

• Lagerklasse: 12 - Non Combustible Liquids
• wgk: WGK 1
• flash_point_f: Not applicable
• flash_point_c: Not applicable