Question

To make cDNA, 400 ng of RNA is required, where the maximum volume of RNA we can add to a 20 uL reaction is 12 uL. As per the Nanodrop spectrum of the sample, it has a concentration of 605.6 ng/uL, A260/A280 ratio of 2.03 and A260/A230 ratio of 1.65.

What is the yield of the RNA? What volume of RNA is required for the cDNA synthesis?

Answer 1

RNA yield: As given in the question, from the nanodrop reading the concentration of RNA sample is 605.6 ng/ ul. This is 605.6 ng/uL is the yield of RNA which means that each microliter (uL) of the sample has 605.6 ng of RNA. Therefore the yield of RNA is 605.6 ng/uL.

The required volume of RNA for cDNA synthesis: The 400 ng of RNA is required for the cDNA synthesis which can be calculated in the following manner with the help of yield of RNA sample:

Since 605.6 ng RNA = 1 uL ( as we know from RNA concentration that each uL (1 uL) contain 605.6 ng of RNA)

So, 400 ng RNA = 1*400/ 605.6 = 0.66 uL

So 0.66 uL volume of RNA is required for cDNA synthesis to set up the reaction.