I am transfecting my DNA into HEK293T cells . The WT cells (which has the plasmid used for cloning) do not show the flag when WB was done , while it was expressing in the mutant i made. Can please someone help with that?
is the Flag fused to something in your WT plasmid? if not only a short protein is make that run out of the gel. does it have a start codon? did somebody used the wt plasmid before? maybe the wt has a mutation that you corrected doing the mutant... did you sequenced the wt and mutant genes?