Difference between DNA and RNA extraction with phenol - DNA or RNA extraction with phenol (Sep/12/2005 )

hi,

what is the main difference between DNA or RNA extraction with phenol. is there any point relating to PH of phenol specific to DNA or RNA


Javaid

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hi
pH is case sensitive :
for RNA extraction : pH is 4.7
for DNA extraction, pH is 8
You can see the ambion's tech notes for further adds (here)
fred

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Hi,

if you use the pH 8 phenol for your RNA extraction, your RNA will be subject to alkaline hydrolysis (it will be degraded).

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hi,

i have phenol from ROTH. The pH is already adjusted with TE buffer and the pH range is 7,5-8,0. this means that i cannot use this phenol for RNA extraction (if i want, then how to adjust the pH of phenol)


javaid

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well you can't use the so mentionned phenol for this purpose...
Did you read my previous post? I gave a link to Ambion's technotes which say how adjust pH of phenol based solutions...
and if it's boring you toadjust pH you can by tri reagent or trizol...
fred

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^
hi,

i read the information, but perhaps there is no informtion for adjusting the ph of phenol. besides this i feel that to work with trizol is easy rather to adjust ph of phenol, any how for knowledge can u refer me information for adjusting pH. and effective protocol of RTPCR for trizol extracted RNA

Javaid

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ouuuuuuuuuuuuuuuups i've putted the wrong reference. Sorry for that... wow. I know that pH of phenol solutions can be monitored by pH paper. But for adjustment, i do not kow. I can't get back the reference... really sorry

i've never done a rtpcr on trizol extracted RNA...
But i assume a DNAse treatment of the RNA is quite a good idea. After that, it's a standard RTPCR...

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To adjust the pH of phenol, you add a top layer of Tris-HCl of the correct pH, shake, allow to separate, and test the pH. If not correct, remove the top water layer and replace with fresh buffer. Repeat as necessary. For acidic solutions (RNA work) you probably want a buffer other than Tris, since its buffering capacity at pH 4.5 is negligible.

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