SLIC

Originally described by Li and Elledge

Cloning by SLIC is an economical strategy for ligating dsDNA. dsDNA is first treated with T4 DNA polymerase which has a 3' exonuclease domain. This creates sticky ends on either end of dsDNA. The reaction is then quenched with the addition of a single nucleotide triphosphate. dsDNA with sticky ends are then mixed together, allowing the sticky ends with significant homology to bind together. This mix is then transformed into \ecoli\ where it is assumed DNA repair machinery repairs the nicks and gaps in DNA mixture are repaired to make a functional plasmid.

Below is my working protocol. There is significant room for optimization on a case by case basis, but the protocol below has been sufficient for most of my cloning. Most of my dsDNA samples have regions of homology with a predicted melting temperature around 65$^{\circ}$C.

SLIC is prefect for ligating two strands of dsDNA together. In such a reaction I would expect about 50-200 colonies on the final plate. One strategy for getting SLIC cloning to work on multiple fragments of DNA is to ligate some of the fragments beforehand via PCR and then using SLIC to create the final plasmid as SLIC is typically faster (45 min)than doing PCR ligation.