what primers should I use for DNA sequencing?? - (Oct/19/2009 )
I only know the vector is pED-3d, but dunno the insertion site. Now I need to submit samples for DNA sequencing, please let me know what primers should I use? (T7? or something else) If I have to make primers myself (without knowing the insertion sites), how should I do it?
Thank you all so so much!!
the insert will normally be in the multiple cloning site (mcs). since you don't know any more then you should use a standard primer (like t7) if it is in the plasmid. try one that is about 100 bases away from the mcs.
mdfenko on Oct 19 2009, 12:40 PM said:
thanks a lot for ur reply!!
vectorpedia has an entry for a mammalian vector pED (~5.4 kb; amp resistant), but nothing for pED-3d. Could it be pET-3d (4.6 kb, amp)??? Do you have a size for the vector? Try some standard digestions to look for unique sites - EcoRI, BamHI, HindIII. You may even pull out your insert...