crRNA & Cas9 Protein

CRISPR/Cas9-mediated gene editing is a powerful technique that allows you to create knock-in/out mutations in any gene and any cell. CRISPR/Cas9 is advantageous over other forms of gene editing, such as TALENs and zinc finger nucleases, because it is simpler to implement and edits at higher efficiency.
 
Our brand partners GenScript licenses CRISPR technology from the Broad Institute of MIT and Harvard. Our offerings include the latest CRISPR plasmids and databases developed by the CRISPR pioneering Feng Zhang laboratory. Broad Institute-validated plasmids are a well-tested platform for expressing CRISPR/Cas9, and avoid instability issues in RNA-based platform
 
PRODUCTS
 
1. CRISPR/Cas9 tracrRNAs
 
CRISPR/Cas9 trans-activating crRNAs (tracrRNAs) are 67 nt RNA oligonucleotides which together with the crRNA and Cas9 nuclease, from the activated CRISPR/Cas9 ribonucleoprotein complex.
 
2. Cas9 Protein
 
Cas9 protein is the CRISPR-associated nuclease which enzymatically cleaves DNA and enables gene editing. Cas9 contains a N-terminal nuclear localization signal (NLS) and C-terminal NLS, for optimized nuclear compartmentalization.
 
3. CRISPR/Cas9 crRNAs
 
CRISPR RNAs (crRNAs) are short 36 nt RNA oligonucleotides which contain a short 20 nt sequence which guide the CRISPR/Cas9 complex to genomic targets for gene editing.

4. CRISPR/Cas9 Control Kit
 
We offer pre-duplexed human HPRT crRNA:tracrRNA as positive controls for your CRISPR experiments. HPRT (hypoxanthine phosphoribosyltransferase) is a housekeeping gene and commonly used control has already been pre-validated for efficient CRISPR cleavage. The kit comes with an HPRT primer mix for PCR analysis.

OVERVIEW
 
Synthetic crRNA/tracrRNA
 
CRISPR/Cas9 ribonucleoprotein (RNP) complexes are composed of a CRISPR RNA (crRNA), trans-activating crRNA (tracrRNA) duplex and Cas9 protein. The crRNA consists of a 20 nt sequence that is complementary to a genomic target. crRNA sequence design utilizes standard gRNA design principles and sequences can be easily selected from the the gRNA database or customized using the gRNA design tool. When complexed with tracrRNA and Cas9 protein, a double strand break will be created at a locus complementary to the 20 nt guide RNA sequence, 3-4 bp upstream from a protospacer adjacent motif (PAM) sequence (5'-NGG-3'). Unlike traditional CRISPR Plasmids, CRISPR/Cas9 ribonucleoproteins are delivered as intact complexes, and do not require cellular expression.
 
 
CRISPR RNA/Cas9 Protein Workflow
 
Only three steps are required for using synthetic CRISPR RNA oligos:
 
1. Incubate the crRNA and tracrRNA with the annealing buffer
2. Incubate the duplexed crRNA:tracrRNA with Cas9 protein to form the active CRISPR/Cas9 ribonucleoprotein. Optionally, an HDR (homology dependent repair) template can also be co-delivered with the Cas9 RNP for targeted knock-in.
3. Deliver the ribonucleoprotein mix into the cells
 
 
Prior to experimentation, we recommend first testing your CRISPR/Cas9 complex in vitro and in test cell lines to ensure cleavage and transfection efficiency. Delivery of the CRISPR/Cas9 complex into cells is typically performed by electroporation or via lipid-mediated transfection.