File:Homologous Recombination Deletion method one.png

Image Explanation

Linear sequences of DNA were created using PCR from synthetic primers (the red stretch of DNA). Regions A, B and C inside the target region inside the genome (green) and inside the linear PCR sequence (red) are homologous regions. Cm^R is antibiotic resistance, S indicates an I-ScsI cleavage site.

The DNA is electroporated into the cell, then a plasmid coding for recombinase elements used for recombination. Cells that succeed in recombination were isolated.

Another plasmid coding for I-SceI meganuclease was inserted into the cell to remove the Cm^R gene. Then, a version of Rec-A mediated DSB recombinational repair was used to ligate the ends together.