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HELP! DNA Quantification

HELP! DNA Quantification - DNA Forum

HELP! DNA Quantification - Discuss DNA, the molecule of hereditary. Topics include DNA structure, DNA replication, DNA function.


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Old 05-09-2011, 07:41 AM
Pipette Filler
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Post HELP! DNA Quantification



Hello,

I had a question about the quantification of DNA using the Nanodrop,

I amplified my DNA using normal PCR amplification i purified it with a Qiaquick PCR purification kit and then i measured my samples using the nanodrop, i had the following results:

Concentration (ng/ul) = 18,0
260/280 nm = 1,78
260/230 nm = 2,88!! <- This value concerns me

Does anyone of you have an idea what this high value means, i have looked up some forums and they said it was because the concentration of the DNA was too low but i don't think that this is the problem

Can anyone help me?
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Old 05-09-2011, 01:30 PM
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Default Re: HELP! DNA Quantification

I would be inclined to agree that it is down to your low DNA conc. Alternatively I have seen the 260/230 value being high when using sureclean and I haven't let the DNA pellet dry properly - so the alcohol interferes. It might be worth running on an agarose gel to check the DNA band quality.
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Old 05-09-2011, 01:44 PM
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Default Re: HELP! DNA Quantification

I ran the purified product on a 2% agarose gel, and i saw no abnormalities...maybe a bit blurry bands but not abnormal..

Could it still be the interference of alcohol?

The only thing i forgot to mention is that the 5' strand of the DNA molecule has a NH2-C6 group attached to it, could it possibly be some kind of absorbance either from the NH2 of the C6 chain?
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Old 05-11-2011, 02:14 PM
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Default Re: HELP! DNA Quantification

I've no experience of adding that group to the end of a PCR product. But I would imagine that it would alter the result in the nano drop. If the DNA bands look fine on agarose then I would be inclined to think that the DNA quality is fine.
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