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<TITLE>Re: [RASMB] AUC of RNA and RNA-proteins complexes</TITLE>
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Dear Pascal -<BR>
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I would give a slightly different emphasis from Jo Butler in replying to your question. Under two headings:<BR>
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(1) <B>value of vbar to be used<BR>
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The value of vbar of RNA <U>as its sodium salt</U> is indeed less than the 0.55 ml/g often used for DNA. But although it is doubtless a bit variable (e.g. may depend on <I>which</I> RNA, precise amount of salt, etc), I think that 0.50 ml/g may be too low a value. For 16S rRNA we long ago measured the value in 200 mM NaCl to be 0.524 ± 0.08 / 0.521 ± 0.007 ml/g for two separate preparations. One of the same samples however, in 200 mM <U>KCl</U> gave a vbar = 0.580 ± 0.004 ml/g ! (the latter value has been found by others). See Pearce, Rowe & Turnock (1975) J Mol Biol <B>97</B> 193-205<B>***</B>.<BR>
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<B>(2) is life hell working with RNA ?<BR>
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As Jo says, no, it need not be, if you take reasonable precautions. I have always found however that 'reasonable precautions' had to go beyond what Jo recommends. RNAse is ubiquitous - your fingertips have it on them, its over your bench, its probably in your glassware and cells, as the wretched stuff recovers even from boiling. The way to keep it at bay is to treat every reagent used with DEPC (freshly added from alcoholic solution, it does not retain activity once in aqueous solution) and to soak/rinse all cells and glassware with distilled water to which DEPC has been freshly added. If all else fails - and on occasion it can do - then adding the (horribly expensive !) 'RNAsein' (a specific pancreatic inhibitor) is called for. <BR>
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What Jo says about the likely range of s value etc I totally agree with. It is probable, by the way, that compact, folded RNA is more resistant to the dreaded RNAse.<BR>
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Good luck !<BR>
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Arthur Rowe<BR>
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<B>***pdf available to be mailed, on request.<BR>
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Arthur J Rowe<BR>
Professor of Biomolecular Technology<BR>
NCMH Business Centre<BR>
University of Nottingham<BR>
School of Biosciences<BR>
Sutton Bonington<BR>
Leicestershire LE12 5RD UK<BR>
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Tel: +44 (0)115 951 6156<BR>
+44 (0)116 271 4502<BR>
Fax: +44 (0)115 951 6157<BR>
email: arthur.rowe@nottingham.ac.uk<BR>
arthur.rowe@connectfree.co.uk (home)<BR>
Web: www.nottingham.ac.uk/ncmh/business<BR>
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