Hi,
I just did a saliva dna extraction and the A260/230 results read “normal”-ish(reading at 1.37-1.9) but almost all samples had low A260/A280 readings. With the exception of one, they read less than 1.
Any idea on what that means and what I should consider for to rectify this?
Hello,
Typical pitfalls:
Carryover of residual phenol or other reagent associated with the extraction protocol
very low concentration of nucleic acid
pH of buffer (acidic gives lower A260/280)f
Although I have never extracted nucleic acids from saliva, it would appear that you have some contamination issues. How exactly are you performing your extractions (e.g. commercial kit, in-house method, etc)? If you're using a personal method, then it's likely something flawed within that. Perhaps a certain buffer/reagent? If you're using a kit, then maybe it's not the best one for saliva...I would suggest trying to find a more optimal kit. I'm certain there are several companies out there selling kits for this type of thing.
Also, if contamination is truly the problem...do not just look at the ratios, but also the actual peaks generated on your spectrum. The size and shape of the peaks can also give insight into potential contaminants. Check the following pdf files I have attach for more information on this.
Also, I would suspect you don't get much DNA from saliva...but I'm not certain on this. But if you are, then low amounts of nucleic acids can also throw off your ratios...even if the sample has "pure" DNA. What are the concentrations you have been getting? Despite poor ratios, you might still have enough DNA to proceed with your downstream applications.
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