I'm a grad student working on metagenomics and I ran into some issues with the samples. I collected urine samples and extracted DNA using DNeasy Blood & Tissue kit. I used NanoDrop to measure DNA concentration and ran into different issues. The problem is that the A260/280 level is too low (not to mention the contamination) and I don't know how to increase the purity level. I don't think I can collect the samples again, or at least it will take a while before I can do that again. Is there a way or a kit I can use to increase the purity level of my DNA extracts without significantly lowering DNA concentration?
Does RNA contamination impact your downstream assay(s)? If no, you may be fine as it is. If yes, you can try RNAse treatment and Zymo has some quite good DNA concentrator kits - but that would still mean ~20% loss or more of the DNA
Péter Gyarmati It's protein contamination, and I believe with a high level of protein contamination, I cannot perform library preparation. I'm already suffering from low DNA concentration so I guess there's not much I can do. Do you have suggestion for purification protocol after extraction that wouldn't substantially lower DNA concentration?
1.If you can, it is possible to isolate the microbes( grow in lab) to increase the number of organisms which indirectly increase concentration as well as purity of DNA.
2. Increase the template during PCR
Sam Kim well, first I would want to make sure it is really protein - nanodrop can give false values for all sort of reagents. Try measuring your DNA w Qubit or Bioanalyzer. If you have no other option but to purify DNA again, Zymo kits are the best in my experience (but you will still lose DNA, I am not aware of a way around that)
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