I-SceI

The I-SceI endonuclease was the first homing endonuclease to be identified and is today the most widely used for research and genome engineering. The I-SceI homing endonuclease belongs to the LAGLIDADG family.

I-SceI was discovered by Professor Bernard Dujon from Institut Pasteur in the 1980s. It is encoded by the Omega intron in Saccharomyces cerevisiae (baker’s yeast) mitochondria. This intron was discovered in the mitochondrial ribosomal RNA of a strain, but not in other strains. When an intronless strain is crossed with intron-containing strain, the intron spreads into the intronless gene copied by a process of gene conversion, with co-conversion of the flanking exons extending to several hundred base pairs upstream and downstream. This conversion is initiated by the endonuclease activity of I-SceI: I-SceI creates a double-strand break (DSB) in the intronless gene copies, and this DSB is repaired by homologous recombination with the intron-containing copy.

The I-Scel DNA target site is an 18-bp sequence. On a random basis, this type of sequence is found in one base pair for every 70 billion (418). Statistically, a genome 20 times the size of the human genome (3 billion base pairs) would therefore be required for there to be a real probability of finding one I-SceI recognition site. This high specificity makes I-SceI the ideal tool for genome engineering in all organisms.

I-SceI has now become a paradigm for meganuclease-induced recombination. It is used as the gold standard to induce recombinations in bacteria, yeast, mammal cells, fish, worms, flies or plants.