What's going on with phenol?

Chadwick, Brad bchadwick at agric.wa.gov.au
Thu Sep 7 05:16:40 EST 2000


Recently the mini-preps I've done using alkaline lysis and phenol extraction
have been giving extremely high OD's, making them look brilliant, until I
check by restriction digest and agarose gel, to find there's very little
plasmid DNA actually in there.  MPs prepared by Qiagen columns worked fine,
but the yield is not good enough for what I want. When I looked into the
problem with the other ones, it turned out that its the phenol. Even pure
water extracted with phenol gives and OD thats off the scale, and it seems
that this contaminant which affects the absorbance at 260nm, is potent
enough to still make a big difference even after the DNA precipitated and
the supernatant poured off.  It happens with my brand new bottle of MB grade
phenol/chloroform/isoamyl alcohol from Sigma and it happens with another
bottle of old oxidized phenol only. I'm certain it never used to happen when
I was doing MPs years ago, before Qiagen columns existed, but I always did
them with phenol. 

If somebody has experienced this before with phenol I'd like to find out
what causes it and whether its avoidable, and particularly whether any
problems were experienced with the DNA in downstream processing, like
ligations etc. 

Brad


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