DNA extraction from Gram positives

THenics at intercell.co.at THenics at intercell.co.at
Thu Aug 19 08:31:50 EST 1999


Hi Elizabeth,

Recently, I developed a relatively quick and neatly reproducible method to
isolate 
genomic DNA from Staphylococcus aureus. Here I first I have to give great
credit 
to Muhammad Sohail (see Molecular Biotechnology, 10: 191-193, 1998) and 
Betley at al (see PNAS, 81: 5179-5183, 1984) whose previous protocols served
as 
a backbone for the one I developed.

Two things which I found perticularly useful are:

1. Both RNase A and RNase T1 are present in the extraction millieu at high 
concentrations (0.5 mg/ml and 10 U/ml, respectively) prior to breaking up
the cell 
wall with Lysostaphin. This I found quite importand to preferentially
eliminate RNA 
contamination.

2. Instead of using a series of PhOH extractions which creates a big mess, I

employed the Strataclean Resin (Stratagene)- which was also suggested
earlier- after 
a single round of organic extraction.

But, drop me an e-mail if you need the full protocol.

Tamas

====================
Tamas Henics, MD., PhD.
INTERCELL, Inc.
Dr Bohr Gasse 7b
A-1030 Vienna, Austria
Phone: 43-1 796 4422/110
Fax: 43-1 796 4422/9
http://www.intercell.co.at
====================




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