I have purified PCR product using PCR purification KIT.
I did a qubit assay to quantify the DNA and yield in the range of 90-110 ng/ul. I tried to dilute it the samples to 13ng/ul - 15ul range (50ul Volume). I did not get the value I was looking for. The error is too high. Some show DNA in the range of 30 ng/ul, some are negative and some of them have the same value as before.
What could be happening? Please suggest a better method to quantify DNA?
Prasanna, are you sure is it not because of pipetting error,? Do you check the volume you pippett before you add into the solution. Do not use small volumes for pipetting such as 1uL or 2uL. In such a case like determination of concentration, its really good to use minimum 5uL volume for pipetting.
Measuring the DNA concentration isn't that reliable... it can just give you an idea
errors are high in measuring at very low concentrations so trust the value that you get at a high concentration where the errors are small then pipette at a volume that gives a low pipetting error and trust the resulting dilutions to have the correct amount of material
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