Gene editing using non-integrating lentiviral vectors

Cre-recombinase was used with LentiFlash in order to evaluate its efficacy in promoting gene excision within target cells stably expressing the dsRed protein from a "floxable" transgene. RNAs transferred by LentiFlash particles are directly expressed and do not integrate after cytoplasmic transfer.

Gene editing experiments in vitro and in vivo with Cre-lox recombination

In vitro experiments

In vitro Cre-lox recombination
In vitro graphe LentiFlash Cre-lox recombination

Following transfer, LentiFlash-Cre particles induced the steady excision of the dsRed transgene (Figure 2, sample 4 for the polyclonal population and sample 7 for clone 14) with high efficiency in all target cells. Of note the stability of excision suggested 100% efficacy following LentiFlash particles mRNA delivery.

Equivalent excision was observed with an IDLV expressing Cre from an EF1 promoter (Figure 2, sample 3 for the polyclonal population and sample 6 for clone 14). The main difference is that unlike IDLV, LentiFlash particles allows the excision without residual integration.

Next, we determined whether LentiFlash enables systemic and localized in vivo expression in mice. The ability to deliver RNA in vivo would be valuable for several research and clinical applications.

In vivo experiments

In vivo Cre-lox recombination
In vivo graphe LentiFlash Cre-lox recombination

The mouse model Figure 4 highlighted the efficiency of the non-integrating lentiviral particle LentiFlash to induce stable excision of the gene of interest (Here YFP).

The excision led to YFP expression in a wide zone of injected muscle (Figure 4, a, b and c, as compared to non-affected muscle zone d). LentiFlash particles spread in approximately 1 cm of the muscle, and this is comparable to the area reached with purified lentiviral vectors.

Altogether, in vivo data confirmed the potency of LentiFlash particles for delivering functional mRNA locally.

Please contact us for your gene editing project with LentiFlash and your gene of interest and fill out the form with the information you are willing to share with us.