Hi everyone, this is something that I´ve been wondering about for a while, it´s just for general knowledge. Does anyone know what´s the minimum concentration of DNA in order to obtain amplified products?
Hi again, thanks for your answer!
My comment was leading to the following problem (which I´m sure many of us has encountered):
I have a DNA sample which is an only sample, and it´s not much (10 microliters) and the concentration is too low (5 ng/ul), therefore I don´t have the opportunity to test it because I could loose the sample.
So I started wondering, how much template should I use to get the amplified fragment on the first try?
Cheers everyone!
Hi everyone, I already amplified the DNA, what I did was to decrease the final volume, so if the PCR didn´t work I wouldn´t loose sample, everything worked up fine.
I just wanted to comment about this because, even though is a simple topic, there are different ideas and other issues to consider, for example: if the sample is an amplicon, purified DNA, plasmidic DNA.
Of course, other aspect to consider is the 260/280 ratio, it´s recomended to be between 1.8 and 2.0
When you quantify your samples, even though you get good DNA concentrations, if the 260/280 ratio is not close to the previously mention values, the amplification doesn´t work (this has been my experience).
Cheers everyone!
yes, walter is right, in colony pcr, one picks up bacterial cells for PCR
Yes, with bacterial colonies there´s no problem
But with purified bands, in my experience, the OD ratio is important.
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