DNA-isolation

Joachim Nerz (joachim.nerz@student.uni-tuebingen.de)
Wed, 17 May 1995 13:47:49 +0200 (MESZ)

Dear Ben,

you asked about DNA from Dionea and Nepenthes;

maybe you are interested in an quite easy protocol for DNA-isolation from
plants;
it's the so called CTAB-method; with that method, I also could isolate
DNA from Nepenthes for an RFLP:

1-3 g leave material grinded to fine powder in liquid nitrogen

add quickly 15 ml 2x CTAB-Buffer and resuspend

(CTAB-Buffer:

100 mM Tris-HCl pH 8.0 1,4 M NaCl 20 mM EDTA 2g CTAB
(Ceytyltrimethylammonium bromide) 0,2% Mercaptoethanole

Incubate 30 - 60 min at 50-60 C (55 C)

add one vol. Chloroform-Isoamylalcohole,

mix by inverting the tube 5-6 times

centrifuge at table-centrifuge (e.g. Heraeus) at 5000 rpm, 15 min, roomtemp.)

take the upper phase and do the extraction again

take the upper phase and add 2 Vol Icecold Ethanol (100%) and put in
refrigerator: -20 C: at least 2 hours
-70 C: at least 30 min

centrifuge 15 min at 4 C, Heraeus 5000 rpm, 15 min

wash the pellet with 70 % Ethanol, again centrifugation 5000 rpm, 10 min

dry the pellet

resuspend it in TE or Aqua bidest. (100 - 500 ul, depending on size of
pellet) (in TE the DNA is more stable)

ready :-).

Good luck

Joe