Since you dont have Mol.biology back ground you should check every step with someone in your lab or at your dept, becos sometimes it might take several months to figure out the problem for freshers.
Check for the same restriction sites like if your DNA is cloned with pCMV6-XL4 at ECORI , NotI and if you have the same Restriction site PCDNA1.1/AMP
all you have to do is digest with these enzyme which will open the pCMV6-XL4, and your Dna will drop out of this vector. Cut the PCDNA1.1/AMP in similar fashion which will open it too. after that gel purify these.
take the dna alone from the gel and purify it, also after enzyme restriction purify the PCDNA then run it on the gel to make sure they are clean, you can check the m.wt. On the gel you should see only a single band, if you have many bands then not enough cleaning
Then go for the ligation
either ligafast promega or NEB overnight ligation both works
after ligation do the transformation, check the control (which should not give colonies) and your ligation, then screen the colonies for the insertion of dna, you can check by doing the same enzyme digestion
It will be more clear if you talk to anyone personally
Edited by ranvi, 28 August 2010 - 01:17 PM.