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Strange nanodrop readings after RNA isolation, strange solution - (Dec/20/2005 )

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QUOTE (John Buckels @ Jan 9 2006, 04:00 PM)
To re-suspend my RNA I actually freeze at -80C overnight then thaw and vortex (rather than risk heating)

PS Your "2nd" round of ratios look fine for both 260/280 and 260/230

Hi John,

I see, so my strange solution is no stranger to you. Great!
But I would like to understand the principle of this method. I can understand that heating dissolves RNA. But why freezing works is a mysterie to me. If we understand how this works, then maybe we can think of a method that is less time-consuming.
Did you already try other methods?

Thanks, Jasper

-Jasper Bok-

your graph seems good but i don't use this software so comments on it will be reduced to minimum... sorry.

here is about an other topic dealing with hith ratios...
"usually this ratio should be between 1.8 and 2 for good DNA quality. i've heard once that for DNA prep, a too high ratio is nefaste and then could be great to do an other phenol chlo.
For RNA, the intervall is quite larger. From 1.6 to more than two(2.3 is still ok). According to the principle of quantitation, a highly pure RNA should have a high ratio.
i don't know what your rna is for, but load part of it on a gel and see how it looks like. take again the OD and include the 260/230 ratio (salts and solvants).
if you want a true OD, it should be taken at 95° where secondary structures of rna are avoided. in fact, OD at room temperature is 10% less than 95°_OD. But assuming that all samples are treated same way, it's not a problem.

if you want to be sure, to an other phenol chlo and precipitate again. but if your ratio is up to 2.2 i don't think it's necessary.


You should avoid air bubble when usin Nanodrop.


Dear Jasper,

Your RNA may not have been dissolved prior to measurement or your sample column may have broken during the measurement giving you the erroneous results the first time. Try using more sample 1.5-2.0 ul since the surface tension of the sample may be lower. Your 260/230 ratio will be very dependent upon your blank. See attachment.



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