Ligation protocol - (Jan/14/2004 )
Have you ever performed a ligation between two inserts and only a vector? I can first ligate one of the inserts and two days after ligate the second one but it would be useful to perform it in a single step.
It is possible to perform a 3-way ligation, but the transformation efficiency for the correct product will be about 10-fold lower than a 2-way ligation. So you need to use competent cells that have relatively high transformation efficiency.