random hexamers as primers for RT in RT-PCR

Robert Hewitt rhewitt at HELIX.NIH.GOV
Mon Mar 6 22:49:20 EST 1995


Someone in my lab has found that random hexamers sometimes work better
than oligo-dT, as primers for RT in RT-PCR.  This seems to be especially
true when the sequence to be amplified is a long way from the 3' end of
the message.  Our explantion is that reverse transcription from an
oligo-dT primer may often not extend as far as a 5' sequence to be
amplified.  From a random hexamer however, it may stand a better chance.  
Please let me know if you disagree with this explanation. 

Can anyone explain what happens when a number of random hexamers anneal to
the same mRNA, so that reverse transcription occurs from a number of
primers along the same message?  In this situation, would the multiple 1st
strand cDNA molecules join up to produce a single molecule?  If this does
happen, I would be grateful if anyone can provide the explanation. 

Many thanks,

Robert Hewitt




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