This method is used for subcloning well defined restriction fragments and is probably not suitable for genomic library construction. The method works for both blunt-end and cohesive termini ligations. Leave a small amount of each band in the gel to show where the DNA migrated then photograph the gel. Wear protective glasses while working with UV light. The concentrations of insert and plasmid DNA given above is an example.
Ligations work effectively even for bands that are barely visible on a gel.
Incubate the ligation mix in a 15 degrees C waterbath for 12-14 hours preferably overnight. Melt the ligation mix on 70 degrees C heatblock for 10 minutes. Store rest of the ligation mix at 4 degrees C for later use. Test pH using a pH paper. Adjust the volume to 1.0 ml.