DiscussionThe genomic integration of foreign DNA is one of the most widely exploited cellular processes in molecular biology. This process employs the two main pathways of DNA recombination in eukaryotes, homology directed repair and the non-homologous insertion of foreign DNA. Typically transgene addition strategies based on homology independent insertion are very efficient in mammalian cells due to the preliminary role of Non Homologous End Joining (NHEJ) in repairing double strand breaks in higher eukaryots. However this process often results in illegitimate integration of introduced sequences therefore posing a risk of unforeseeable genomic alterations.
Here we show that NHEJ pathway can be used for the precise integration of foreign DNA sequences into the site of breakage generated by custom design nucleases (ZFN, TALEN and CRISPR). In particular we describe a method that we named ObLiGaRe that facilitate the fast generation of models of disease but also the generation of stable cell lines for protein expression. Finally we will provide some examples of the use of precise genome editing in drug discovery.