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dna purity is 17? what does it mean?

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dna purity is 17? what does it mean?

Postby genetherapy » Thu Jan 26, 2012 5:05 pm

Hi everyone,
I'm constructing a new vector and before starting ligation I measured dna purity of my gene. The dna purity that I measured is 17. More more than 1.8 :? What does this mean, is my dna is so pure from proteins etc :D I measured this three times, it is unable to understand for me. Could anyone explain me this?
Thanks for yor help
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Postby JackBean » Thu Jan 26, 2012 5:35 pm

are you sure it's 17 and not 1.7? How did you calculate that?
http://www.biolib.cz/en/main/

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Re: dna purity is 17? what does it mean?

Postby genetherapy » Fri Jan 27, 2012 6:48 am

Yes I'm sure that it is 17 not 1.7. The spectrophotometer calculates automaticly when I enter my dilution ratio. I used spectro lots of times and that is the first time ı see a value like this. But ı make dna cleaning and concentrating prosedure and forget my dna sample 5 or 7 minutes in the final step(elution step). Is this the matter I really don't undestand??
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Postby genetherapy » Fri Jan 27, 2012 6:49 am

I mean ı forgot my sample to centrifuge for about 5 or 7 minutes besides only 1 minute waiting.
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Postby JackBean » Fri Jan 27, 2012 8:21 am

you're centrifuging it for 5 minutes?

Anyway, what was your concentration? Such out-of-mind values usually come up when you have very low concentration and thus very low absorbance.

BTW with the 260/280 ratio higher than 2, the DNA is contaminated with some RNA, phenols or something like that. But I doubt anyone ever saw 17 :lol:
http://www.biolib.cz/en/main/

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Re: dna purity is 17? what does it mean?

Postby genetherapy » Fri Jan 27, 2012 3:48 pm

Interestingly my dna concentration is 157,07 mikrogram/ml :shock:
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Postby genetherapy » Fri Jan 27, 2012 4:04 pm

I also want to ask another question like this problem. Today I purified DNA from pellets. Then I read their purity and concentrations in spectrophotometer. The purities range between 1,8-1,9. Really good and concentrations are good also. Then I digest this samples with an enzyme and look if it digest, the digestion was perfect. I cut the bands that I want and the problem starts at this point: I made gel extraction and now the purities range between 3,5-5 . The concentrations are 5 mikrogram/ml. I'm so unhappy that you can't guess after the perfect purity and concentration from first step. Is it the agarose that wastes my samples? What will I do?
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