I had the same problem. First I cut my plasmid with different enzymes
which produce 3'-overhang ends. Making run-off-transcription with
T7-RNA-polymerase I always got a smear from the size of my expected
fragment up to the size of the whole plasmid.
Then I tried to cut my plasmid with an enzyme which produces 5'-overhang
ends. Now my transcription produced one single band.
Poopak Banky schrieb:
> Question; i was wondering if anyone has encountered variability in RNA
> transcription products when using linearized plasmid templates?