Streptococcus pyogenes Cas9 (CRISPR associated protein 9) is a 160kD RNA guided endonuclease that introduces site specific cleavage of double strand DNA (1).
It is part of the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) system found in many bacteria such as S.
pyogenes and most archaea, which provide adaptive immunity against invading mobile genetic elements (such as viruses, transposable elements and conjugative plasmids) (2, 3).
Upon viral infection, short viral DNA (known as "spacers") integrate into the host genome between CRISPR repeats, and RNA sequences (guide RNA or gRNA) with this genetic information help guide Cas9 protein to recognize and cut foreign DNA.
Cas9 protein undergoes conformational changes upon gRNA binding that shift from non-DNA binding conformation into an active DNA binding conformation.
In the Cas9-gRNA complex, the gRNA sequence remains accessible to interact with free DNA, and the extent to which the gRNA spacer and target DNA segment (known as "protospacer") match will determine the cut site (4).
The presence of a 5′-NGG-3′ protospacer adjacent motif (PAM) sequence immediately downstream of protospacers is required for Cas9 cleavage of the foreign DNA.
PAM is absent in bacterial CRISPR loci, therefore preventing cleavage of the host genome (4).
Cas9 associates with other proteins of the acquisition machinery (Cas1, Cas2 and Csn2), presumably to provide PAM specificity to this process (5).
This RNA guided nuclease system called CRISPR/Cas (CRISPR associated protein) has been widely applied to genome engineering with increased efficiency (6).
The attached nuclear localization signals(NLSs) on the chimeric protein ensures nuclear compartmentalization in mammalian cells during gene editing (7).
Applications:Suitable for use in Western Blot.
Other applications not tested.
Recommended Dilution:Western Blot: 2ug/mlOptimal dilutions to be determined by the researcher.
Storage and Stability:Lyophilized and reconstituted products are stable for 12 months after receipt at -20℃.
Reconstitute with sterile PBS.
Aliquot to avoid repeated freezing and thawing.
Store at -20℃.
For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
Further dilutions can be made in assay buffer.
仕様
Size：100ug
抗体種類：Mab
宿主：mouse
Grade：Affinity Purified
Purity：Purified by Protein A affinity chromatography from hybridoma culture supernatant.
Form：Supplied as a lyophilized powder in PBS, 5% trehalose. Reconstitute with 200ul sterile PBS.
