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Applications: Generate a SaCas9 over-expressing cell line for knockout or knock-in studies.
Description: Cas9 is an endonuclease enzyme that introduces a double stranded break into the DNA when recruited to a specific DNA sequence by the sgRNA (single guide RNA). This double stranded break is repaired in mammalian cells either through Non-Homologous End Joining or Homologous Recombination. Non-Homologous End Joining often results in the deletion or insertion of several base pairs at the cut site, which, when resulting in a frameshift, causes the functional inactivation of the gene.SaCas9 (Staphylococcus aureus CRISPR-associated protein 9) has demonstrated high cutting efficiency in mammalian cells, and its smaller size makes it ideal for packaging into AAV. SaCas9 recognizes a longer protospacer adjacent motif (PAM) site, 5'-NNGRRT-3', than the more traditional SpCas9 (Streptococcus pyogenes CRISPR-associated protein 9). These AAV particles constitutively express SaCas9 under the control of a CMV promoter.Adeno-Associated Virus-DJ (AAV-DJ) is a synthetic serotype made from eight different wild-type AAV serotypes (AAV2, 4, 5, 8, 9, avian, bovine, and goat AAV) using DNA shuffling. These modifications give the AAV-DJ serotype improved transduction efficiency in vitro and in vivo compared to wild-type serotypes. Consequently, AAV-DJ can infect a broad range of cell types.
Formulation: AAV-DJ was produced in HEK293-AAV cells and is supplied in PBS-MK (PBS Magnesium-Potassium) buffer containing 0.01% Pluronic F68.
Purification: The purity of the AAV particles was confirmed to be greater than 90% by staining with One-Step Lumitein™ UV Protein Gel Stain (Biotium #21005-1L). The purity will vary with each lot; the exact value will be provided with each shipment.
Storage Stability: AAV is shipped with dry ice. For long-term storage, it is recommended to store AAV at -80°C. Avoid repeated freeze-thaw cycles. Titers can drop significantly with each freeze-thaw cycle.
Supplied As: Two vials (50 µl x 2) of AAV at a titer ≥1 x 1012 TU/ml. The titer is determined by qPCR and will vary with each lot; the exact value will be provided with each shipment.
Warnings: Avoid freeze/thaw cycles
Biosafety Level: BSL-1
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