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Are you looking for a reliable and affordable way to knockin a gene? The CRISPR Cas9 system is the tool of the century for accomplishing targeted gene knockins.
In this video, the CRISPR Cas9 gene editing system is used to knockin the RFP gene into the AAVS1 Safe Harbor site in HEK Cells. The Safe Harbor site is a preferred site for gene knockins as it enables robust transgene expression with minimal toxic effects. First, we design an All-in-One CRISPR Cas9 sgRNA vector construct with the sgRNA targeted to the human AAVS1 locus. A donor plasmid containing the template for homology directed repair is also designed. After successful transfection, the success of the CRISPR Cas9 Knockin is verified using a PCR-based technique. This technique utilizes custom designed primers targeting a site within the RFP knockin. Knockin is confirmed when a PCR product is observed.
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