I get band double the size what does it mean - (Jul/17/2013 )
hello guys I am new to this forum
I have a problem, I transformed my gene into pichia using pPICZalpha vector , i found that the yeast is growing on zeocin plates n i confirmed that my vector has been inserted into it
to confirm the transformation I ran PCR usin 3' n 5' primers along with a control.
I expect to get a band of same size as that of the control but somehow i see a band which is double the size of the control band
my question is what that implies
It could mean that you have a double copy of the gene in the plasmid, or it could mean that your PCR is non-specific (i.e. there are other binding site in the vector), or it could mean that the sequence you thought was in there is actually something else.
Were the primers specific for the insert or were they off the vector?
bob1 on Wed Jul 17 21:32:13 2013 said:
It could mean that you have a double copy of the gene in the plasmid, or it could mean that your PCR is non-specific (i.e. there are other binding site in the vector), or it could mean that the sequence you thought was in there is actually something else.
Were the primers specific for the insert or were they off the vector?
the primers were specific for the vector
Ok, have you tried getting the product sequenced then?