cDNA synthesis primer - (Jun/23/2006 )
I found different lab use different primers for cDNA synthesis. My lab use random decamer. Another lab uses radom hexamer. The third lab uses oligo dT primer? Which one is better? Or they gonna do the same thing?
Since only the mRNA (and very few other RNA) has a plyA tail, if you use the oligodT you will basicalli amplify the mRNA only. If you need to use this RNA for real-time PCR and your control is another mRNA, it's fine. If you want to use a rRNA, then it's not the best choise I think. Using random hexamer, you will not discriminate between the mRNA you reverse transcribe. I've never used the random decamer but I guess they work like the random hexamer
Of course, using oligo-dT primers you are running the risk of getting a bias toward 3'-located sequenced, as cDNA synthesis will start at the 3' end of your mRNA (and as was said before, you may lose information about non-polyA RNAs).
You might want a look at this article:
Reverse transcription using random pentadecamer primers increases yield and quality of resulting cDNA
Michael Stangegaard, Inge Høgh Dufva, and Martin Dufva
BioTechniques Vol. 40, No. 5: pp 649-657 (May 2006)