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You are in DNA (PCR, Real Time PCR) Forum >> Assay Development Sub-forum

High 260/230 ratio for DNA

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Kristina M.
Croatia

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Topic Started by Kristina M.
on 7/3/2009 1:22 AM
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I purified my DNA samples form whole blood using QIAamp Blood Maxi Kit (Spin Protocol). Using Nanodrop, my 260/280 ratio was ok (around 1.8), but 260/230 ratio in some samples was as high as 8! I know that ideal 260/230 ratio is around 2, and if it is lower that probably means contamination with organic solvents. Does anybody  knows what  high ratio means?


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Shampa
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Posted By Shampa on 7/3/2009 2:58 AM
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 You are right it should be aroung 1.5 ideally. What I know from my previous experience is that the high ratio indicates high salt concentration in your sample. Please try to reduce it.



Jiten
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Posted By Jiten on 7/3/2009 16:41 PM
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I may be wrong also. Shouldn't we expect the A260 / A230 ratio to be low if the salt concentration is high in the samples? I have experienced this problem and I have not been able to sort out what the error happening with similar A260 / A230 ratios. I have got even once at 18.



sps
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Posted By sps on 7/3/2009 3:30 AM
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I would advice diluting your sample in 10mM Tris-HCl at pH 7.4 but not water. One more thing, low ratios may be due to phenol contamination. Check it out!!! High ratios might come from salts in yoour samples. I never saw ratios as high as indicated here. But if you used Trizol, maybe try doing clean up using a column spin methods and try again. Wish you good luck. 

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Monu
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Posted By Monu on 7/3/2009 3:54 AM
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 I also suspect the high salts conc is the main reason behind it. But you should check spec instrument functioning also.

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LBurger
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Posted By LBurger on 11/9/2009 10:18 AM
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 Wouldn't a very high 260/230 ratio indicate a low (in in the case of some samples I just OD'd zero) absorbance at A230. Personally, I'm more concerned with contamination issues when I have significant absorbance at A230 indicating phenol, salt, etc (and low 260/230) than when it's too high. From what I've read anything over 2 is good.

Low A230, could also be related to how dilute your sample is; if the A260 is low (say 0.05), then it may be difficult to accurately estimate A230 resulting in over inflated 260/230 ratios. 

I'd remeasure the samples more concentrated, if possible, and see if you can get a better A230 reading. I like to shoot for an A260 of ~0.1.  However, as I said above I think a higher ratio (low A230) is better than low.



Kristina M.
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Posted By Kristina M. on 11/12/2009 2:56 AM
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After concentrating my samples (using ethanol precipitation after adition of 3M sodium acetate) and re-measuring the concentration, my A260/230 ratio was normal! I suppose the reason for the high ratio could be high initial salt concentration, but whatever it was it is gone after this procedure



Susie Q
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Posted By Susie Q on 12/13/2009 16:56 PM
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I`m working with AFLPs, so I need a nice DNA that could be completely digested. I have seen that DNAs with low 260/230 ratios don`t have fully digestion... it is logical... but also DNAs with High 260/230 ratios (more than 4) just parcially digest. I am wondering myself why this could be happening????





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