Recombination-independent genome editing through CRISPR/Cas9-enhanced TargeTron delivery

Group II introns are retroelements (elements that are transcribed into RNA, reverse-transcribed into DNA and then inserted into a new site in the genome) that are found in bacterial and organelle genomes and that, when in RNA form, are folded into conserved secondary structures. These secondary structures contain six domains, five of which are very similar among group II introns, whereas domain IV diverges in length between different introns. This is because, in some cases, domain IV includes includes an open reading frame that is translated into a protein used for the mobility of group II introns. Specifically, this this protein is involved in scanning the DNA molecules to find a suitable insertion place, and then participates in the integration process. These introns always tend to integrate into the same places, and the first biotechnological application took place when group II introns were modified to insert into different places of the genome (i.e. TargeTron, that uses algorithms to predict good insertion points, and indicates primers to modify the molecules through PCR). These tools can be used for the generation of knockout mutants or as delivery systems. Nevertheless, they have some limitations, such as the changing integration efficiency which can difficult identification of the mutants, and limitations in the length of sequences that domain IV can harbour.

The present work focuses on overcoming these limitations. How? By combining group II introns with CRISPR technology. Read more in the full article available online here.

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