While CRISPR/Cas9 mediated gene editing is a powerful technique for genome manipulation, two significant challenges remain: obtaining efficient delivery of Cas9 and the gene-specific single guide RNA (sgRNA) to all cell types, and achieving fewer off‑target effects. To address these challenges, we have developed cell‑derived nanovesicles called gesicles that are created using the Guide-it CRISPR/Cas9 Gesicle Production System. CRISPR/Cas9 gesicles contain active Cas9 protein complexed with a sgRNA specific to a gene of interest, and mediate binding and fusion with the membrane of a wide range of target cells. These features can increase gene editing efficiency to levels that are often higher than those obtained using plasmid‑based delivery methods. Additionally, this method allows for tight control of the dose and duration of the Cas9‑sgRNA complex in the cell, with the added benefit of reduced off‑target effects. The nanovesicles are produced in a Gesicle Producer 293T Cell Line via co-overexpression of packaging mix components, including a nanovesicle-inducing glycoprotein and a protein for cell surface display that mediates binding and fusion with the cellular membrane of target cells. Simultaneous overexpression of the Cas9 protein from Streptococcus pyogenes and a target-specific guide RNA (sgRNA) results in incorporation of the Cas9/sgRNA complex within the gesicles. After the resulting Cas9/sgRNA gesicles are harvested and applied to your target cells, they will efficiently enter the cells and mediate efficient gene editing.
This system provides the components needed for cloning and expressing your target-specific guide RNA, and packaging reagents for producing CRISPR/Cas9 gesicles. The Gesicle Producer 293T Cell Line is sold separately.