I want to set up PCR for 16s rDNA in bacteria. At least how many ng/ul should the nanodrop measurement be as a result of DNA isolation?
Dear Berfin, nanodrop can safely measure the lowest possible concentration of 0.4 ng / ul. But for PCR I would highly recommend to use at least 10 ng / ul in total starting template provided that your samples OD 260/280 and 260/230 ratios both are within their normal ranges to confirm the purity and quality of your extraction. I usually perform conventional PCRs and qPCR both with at least
25 ng / ul of sample, and if ever I faced any problem it is mainly related to primers not the template (DNA / cDNA) itself.
Regards....
I would recommend 10ul in total to get most visible results for you, I have always used at least 10ul for the conventional PCR. My bands were on good track after doing with 10ul. If you have less concentration then use the PCR result as load for another round to get Quality and Quantity.
brace Berfin Eroğlu
50ng as starting concentration does give results with PCR as well as rtPCR
Berfin Eroğlu
ng/uL is a concentration unit. You should've asked 'how many ng you should use.'
If your nanodrop gave you the DNA concentration is 100 ng/uL, and a PCR reaction needs 50 ng, then you should take 0.5 uL of the DNA (100 ng/uL x 0.5 uL = 50 ng) in a PCR tube for reaction.
Nanodot gives u result in µg/ml . convert µg to nanogram. 1µg= 1000ng.
so accordingly 50ng will be adquate.
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