Frequently Asked Questions: cDNA synthesis primers

cDNA Frequently Asked Questions (FAQs)

Question: Which primer type should I use for my cDNA synthesis?

Answer: This very much depends on your sample type. In general for cDNA generation from full length mRNA you can use oligo (dT) primers, however oligo (dT) primers cannot be used for RNA that does not have poly A sequence, so in this case you can use random primers which allow for the synthesis of cDNA from all RNA in the sample.

Also because of potential 3’ bias with oligo (dT) primers, random primers may be a better option with poly A mRNA when the downstream PCR primers are designed to the 5’ end of long RNA.

One additional consideration is that some mRNAs can have a higher cDNA yield with random primers whereas other mRNAs may have higher cDNA yields with oligo (dT) primers, so optimising the reaction by testing both priming options would be best. However to save time (and reagents) you can simply use a combination of random and oligo (dT) primers.

Finally just an extra note, for 1-step RT-PCR experiments gene-specific primers are the only priming option.

For more information on primer selection see the image below: A comparison of cDNA synthesis primer options.

For a free sample of our FIREScript cDNA synthesis mix or kit and to enter our competition to win a JBL FLIP-4 Portable Bluetooth Speaker... scroll down!

FIREScript Reverse Transcriptase

Fast: 15 minute cDNA synthesis

Stable: Stable at room temp (so "ice-free" set up)

Wide working range: 37-60 degrees C

High specificity and Yield

Available as both a cDNA synthesis mix (with premixed dNTPs and range of primer options) and a cDNA synthesis kit (with everything in separate tubes). Follow the link above to see all available options.

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