CRISPR01

Sigma-Aldrich

CRISPR Human EMX1 Positive Control

NACRES:
NA.51

Quality Level

packaging

pkg of 2 vials (50μL aliquot for each of the 2 kit components)

concentration

20 ng/μL in TE buffer; DNA (1μg of plasmid DNA)

application(s)

CRISPR: suitable

shipped in

dry ice

storage temp.

−20°C

General description

Validated CRISPR site, which serves as an experimental control for the Wt Cas9. A two component positive control system consisting of a CMV-driven Cas9 plasmid and a U6-driven guide RNA plasmid targeting the human EMX1 gene.

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

Features and Benefits

Serves as an experimental control for the CRISPR editing workflow using WT Cas9. Allows for validation of your system with the CRISPR/Cas9 system. A positive result in a miss-match detection assay will indicate validation of your system.

Components

1 vial containing 1ug of U6-gRNA plasmid expressing a guide sequence to human EMX1. 1 vial containing 1ug of Cas9 plasmid.

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). 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.

Physical form

Sigma U6-gRNA plasmid expressing a guide sequence to human EMX1 supplied at a concentration of 20ng/ul in 50ul. Sigma Cas9 plasmid 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

Typical transfection concentrations used in literature are in the ranges of >= 1.0ug/uL and <= 5uL of Cas9 plasmid combined with >= 1.0ug/uL and <= 5uL of U6-gRNA plasmids. (All dosages above assume 0.5 to 1 million cells nucleofected)

storage_class_code

12 - Non Combustible Liquids

WGK Germany

WGK 1

Flash Point F

Not applicable

Flash Point C

Not applicable

Certificate of Analysis

Certificate of Origin

Articles
CRISPR endonucleases have shown wide variation in their activity, even among multiple CRISPRs designed within close genomic proximity.
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View experimental data showing crispr/cas expression and enrichment using FACS
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Protocols
Learn about CRISPR Cas9, what it is and how it works. CRISPR is a new, affordable genome editing tool enabling access to genome editing for all.
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Related Content
Sigma-Aldrich® Advanced Genomics is the leading provider of gene editing and silencing technologies including CRISPR, Cas9, synthetic guide RNA (sgRNA), and Zinc Finger Nuclease (ZFN).
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