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This might be a very stupid question, but I am worried about RNase/DNase contamination of my samples. Since I use 37% HCl and 10M NaOH to pH almost all my buffers, this is a potential source.

I have read however, that both NaOH and HCl should hydrolyze the peptide bonds in amino acids at high concentrations. For HCl, however, I have read that a heating step is required to facilitate the protein hydrolysis, so am a bit confused. Perhaps someone with a better background in chemistry could help?

Thanks a lot!

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  • $\begingroup$ I could just guess it could be safe, because I have not seen the data suggesting it is OK. NaOH or KOH is used to wash glass ware because it has excellent cleaning effect. You could wash away even if you have some RNase in NaHO. But you might miss something making more trouble than that. How about the electrode of pH meter. So, it is good idea to make efforts to remove RNase activities after making buffer. If you really want to RNase free reagents, check if any companies are selling Nuclease free products. $\endgroup$ – 243 Nov 23 '15 at 21:05
  • $\begingroup$ RNAse-A is a very stable protein and tolerant to alkaline conditions. It is used in alkaline lysis and remains unaffected by moderately high pH (0.2N NaOH- Solution II). It even survives GTC and GnCl treatment (refolds when the denaturant is removed). I think 10M NaOH is a very high concentration and the acid/alkali stocks are certainly not a source of RNAse contamination. Pipette tips are one place to look for contamination. $\endgroup$ – WYSIWYG Nov 24 '15 at 3:37
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You are safe here. RNase is much more stable than DNase (of any flavor). Extreme pH would destroy even RNase, especially considering time (it has to sit in the bottle with all that acid/base..) Also, consider this: it is highly unlikely that high-tonnage chemicals like HCL or NaOH could be contaminated with organic enzymes. If you have a problem with RNase/DNase contamination I would look elsewhere for the source of it.

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Treatment with NaOH is one of the methods used to prepare equipment that needs to be RNAse free. DEPC is preferred in my experience, but NaOH certainly works as well. Highly concentrated NaOH like in your case will certainly destroy any RNAses present.

I'd assume the same for concentrated HCl. RNAse is very stable for a protein, but concentrated HCl is not something I'd expect it to survive.

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  • $\begingroup$ RNAse-A at least survives 0.1N NaOH (in solution-I of alkaline lysis 1:1 with solution-II containing 0.2N NaOH) $\endgroup$ – WYSIWYG Nov 24 '15 at 7:02
  • $\begingroup$ @WYSIWYG Interesting. There are quite a few protocols I've seen that use relatively low NaOH concentrations over night to remove RNases, though the one I used myself used 2M NaOH and worked for me. $\endgroup$ – Mad Scientist Nov 24 '15 at 7:06

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