FAQ: CRISPR - Polyplus

CRISPR mediated gene editing requires the delivery of the guide RNA and Cas9 protein into target cells using transfection. The validated method is based on the delivery of guide RNA and Cas9 protein as plasmid DNA, for which we recommend our latest technology jetOPTIMUS®. jetOPTIMUS® ensures highest gene editing efficiency in the largest range of primary cells and cell lines.

DNA-free gene editing methods are considered when off-target effects using the traditional plasmid-based method is a concern. Delivery of gRNA and Cas9 protein as mRNA is the most frequently used DNA-free alternative to reach robust gene editing efficiency in both adherent and suspension cells.

For a full RNA approach – mRNA encoding Cas9 and guide RNAs – we recommend our jetMESSENGER® transfection reagent.

Lastly, for a mix approach – plasmid DNA encoding Cas9 and gRNA as RNA- the most adapted reagent would be jetPRIME®.

For in vivo CRISPR mediated gene editing, our reagent of choice is in vivo-jetPEI®. Several groups have published their results using in vivo-jetPEI® for gene editing in mice:

Zuckermann, M. & al. (2015). Somatic CRISPR/Cas9-mediated tumour suppressor disruption enables versatile brain tumour modelling. Nat Commun 6, 7391

He, S. & al.  (2016) Annexin A2 Modulates ROS and Impacts Inflammatory Response via IL-17 Signaling in Polymicrobial Sepsis Mice. PLoS Pathog 12, e1005743

Takao T. et al. (2020) Optogenetic regulation of embryo implantation in mice using photoactivatable CRISPR-Cas9. Proc Natl Acad Sci U S A 117 28579-28581

Pallarès-Masmitjà M. et al. (2021) Find and cut-and-transfer (FiCAT) mammalian genome engineering. Nat Commun 12 7071

McFadden T. et al. (2022) Short-term exposure to an obesogenic diet causes dynamic dysregulation of proteasome-mediated protein degradation in the hypothalamus of female rats. Nutr Neurosci 1-13