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can T7/SP6 transcribe eukaryotic DNA without bias

silin zhong via methods%40net.bio.net (by sz2840 from gmail.com)
Fri Sep 25 23:21:12 EST 2009


Hi

I am wondering whether one can convert large eukaryotic genomic DNA  
fragments to RNA. It is relatively easy to generate a promoter  
sequence, since adaptor containing T7 sequence (or other types of seq)  
can be easily ligated to gDNA fragments. Alternatively, PCR primers  
containing such promoter sequence can be used. I am not sure whether  
DNA regions with some secondary structure mimicking a T7/SP6  
terminator will cause premature stop of the transcription hence no  
full-length transcript.

If the RNA polymerases do have such a bias, genomic amplification kits  
with a DNA->RNA step will be problematic.

cheers
silin



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