Which strand does T7 RNA pol transcribe from?

brett brett at BORCIM.WUSTL.EDU
Mon Mar 10 09:37:55 EST 1997


>A simple enough question,  but one which is causing me some bother! 
>
> I have a double stranded DNA molecule about 150 bases in length.
>The problem is that the two strands are only complimentary for the
>first 60 or so bases.   At the 5' end of the sense strand I have a T7
>RNA Pol promoter sequence.  My question is: upon introduction of T7
>RNA Pol which strand is the template for the RNA Pol - the strand
>containing the promoter sequence or the strand containing the
>complement of the promoter sequence? 
>
> My understanding of the situation is that the RNA pol uses the
>antisense strand as the template (i.e. the strand with the sequence
>complimentary to the promoter sequence). Therefore the rna generated
>will  be the compliment of the antisense strand.   Therefore when the
>sequences are no longer complimentary (after about 60 bases) the rna
>generated will continue to be the compliement of the antisense strand
>and will include the sequence unique to the antisense strand.
>
>Am i correct?
>
>Any help is appreciated.
>
>gordon
>

You are correct. In fact, check out: Milligan, JF and Uhlenbehck, OC Methods
in Enzymology, V180, pp180-62. here they demonstrate transcription off the
follwoing template:
     5'-TAATACGACTCACTATA-3'
     3'-ATTATGCTGAGTGATATCCCTGTACTCCTAATGGGTACA-5'
                         --------------------->


Brett Lindenbach
    
Program in Immunology                              
Washington University - St Louis                  
brett at borcim.wustl.edu                             




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