Originally Posted by florenciatell
I would like to know in what situation I must use a linker with phosphates for making a ligation to a vector with T4 ligase. Becouse of all the DNAs have phosphates on the extremes, why I should ask for synthesize my oligo with phosphates?
If your vector is cut with 2 different enzymes, you don't need phospho-oligos, because the vector has 5' phosphates.
Instead of ordering the oligos with phosphates (which I think is a bit pricey) you can just do a kinase reaction (Polynucleotide Kinase) to put on 5' phosphates.
But as I said if you make oligos with overhangs that match your vector and use 2 different restricions sites, then you don't need the oligo to be phosphorylated.