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neale at mbcf.stjude.org neale at mbcf.stjude.org
Mon Aug 23 15:08:03 EST 1993


In article <CBoxu9.G0t at biobase.aau.dk>, nelleman at biobase.aau.dk (Lars Nellemann) writes:
> Guanidin-thiocyanat acid phenol vs. CsCl centrifugation in RNA
> purification.
> 
> Has anyone observed that large or small transcripts are selectively 
> removed by using one of these methods.
> 
> I have observed, that small RNAs as tRNA are removed during CsCl
> centrifugation, but not when using the guanidin thiocyanat-acid phenol
> purification.
> Has anyone observed the same, and is it possible that the reverse
> is possible, that very large RNAs are removed during acid-phenol but not 
> in CsCl centrifugation.
> 
> Please reply at:
> 
> nelleman at biobase.aau.dk
>  
>  

We have noticed differences between GIC-CsCl purified RNA and that by GIC-acid
phenol, but have been at a loss to understand why. It seems for us that the
yield of high MW RNA is severely reduced using the phenol extraction method,
and we are wondering if nucleotides and small oligos are contributing to UV
absorbance to the extent of giving a falsely high spectrophotometric reading.
The end result is that 20 ug of RNA purified both ways looks vastly different
on EtBr-stained gels (with concordant probe signal intensity).

For us, for now, when we want the best RNA prep with reproducible yields we go
to the GIC-CsCl method. (It also allows us to collect the DNA at the same time
when we need to)

Regards,

Geoff NEale



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