Overview of the XS DNA extraction protocol for polysaccharide rich organisms
This method is based on the use of the polysaccharide solubilizing chenical xanthogenate. The method
is non-toxic and requires no enzymatic or mechanical cell breaking steps. It has been used with a wide
range of microorganisms, including fungi, gram positive
and negative bacteria, and archea, as well as from plant and animal tissue. The method also works
well with environmental samples like cyanobacterial blooms
and soil. The protocol was published in Tillett, D. & B. A. Neilan (2000) Xanthogenate nucleic
acid isolation from cultured and environmental Cyanobacteria.Journal of Phycology 36: 251-258.
XS buffer
0.5 g Potassium ethyl Xanthogenate (Fluka, 140-89-6)
10 mL 4M Ammonium acetate
5 mL 1M Tris-HCl pH 7.4
2 mL 0.45M EDTA
2.5 mL 20% SDS
Water to 50 mL
DNA extraction protocol
Pellet 1 – 2 mL of cell culture (~100 mg wet weight) in a 2 ml eppendorf tube.
Remove supernatant and resuspend cell pellet in 2 ml of XS buffer.
Incubate at 65?C for 2 h.
Vortex tube for 10 s then incubate on ice for 30 min.
Centrifuge at 14 000 g for 10 minutes
Carefully remove supernatant to a new tube and add 1 volume of 100% isopropanol.
Incubate at room temp for 5 min.
Centrifuge 14 000 g 10 min.
Wash pellet once with 70% ethanol.
Resuspend pellet in 200 µl of 10mM Tris/1mM EDTA (pH8.0) + RNaseA (optional).