HELP: Ligation Problems - (Jun/14/2007 )

First you need to ascertain with certainty
1- that your insert is really in the plasmid (perhaps by sequencing data or PCR data)
2- the vector is correct. Do a digest that cleaves the vector into multiple fragments to make sure it is right

If you are certain of (1) and (2) and have done all you can, then it is time to change ligation strategy. Sometimes a stratergy just don't work for some insane reason.

How about this
Try moving your insert into a pBS backbone or similar (something that has colour testing). Make alot of insert. Then, try to move it into the vector.