Hello,
I used pMES (5.5kb) as a vector and an insert is 1.5kb. My polylinker corresponds to XhoI restriction sites and the sequencing results showed successful insertion of the oligos. I also de-phosphorylated the vector before trying to insert the DNA.
Although colony PCR showed positive results , sequencing results showed no insert. What might went wrong during the cloning that the vector closses itself without an insert?
Thank you in advance.