AAV5 SaCas9
Adeno-Associated Virus Serotype 5 (AAV5) differs from other parvovirus serotypes according to serological and DNA hybridization data, and AAV5 also uses different inverted terminal repeats (ITRs) compared to other AAV serotypes. AAV5 is the most efficient vector for transducing sensory neurons and is good at mediating gene transfer into human and murine airway epithelia. In addition, AAV5 vectors show a higher tropism for both mouse and human dendritic cells than AAV1, AAV2, AAV7, or AAV8.
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.
AAV was produced in HEK293-AAV cells and is supplied in PBS-MK (PBS Magnesium-Potassium) buffer containing 0.01% Pluronic F68.
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