Genomic DNA Extration:
1. Clip a small piece of tail and place in a 1.5ml tube.
2. Add 500ul of tail lysis buffer (100mM NaCl, 10mM Tris-HCL pH7.5, 50mM EDTA
pH8.0, 1% SDS) and 12.5ul of Proteinase K (20mg/ml).
3. Incubate tubes at 56C overnight.
4. Add 500ul of phenol/chloroform to each tube, and gently mix several times.
5. Pour the sample into a phase lock gel tube.
6. Spin tubes at 13,200rpm for 10 mins.
7. Transfer upper layer to a fresh tube.
8. Add 1ml 100% ETOH, and gently mix several times. (A white, stringy material
should coalesce.)
9. Spin tubes at 13,200rpm for 10 mins.
10. Pour off the supernatant and wash the pellet twice with 400ul of 70% ETOH. (Do
not lose the pellet!).
11. Carefully invert the tubes and let them air dry for 10-15 mins.
12. Add 100ul of 1x TE buffer.
13. Incubate the tubes at 56C for 10-15mins. Flick the tubes to resuspend the sample, if
the pellet does not go into solution try incubating the tubes at 37C overnight.
14. Use 0.5ul in your genotyping PCR reactions. (The remainder can be stored at -20C if
needed.)
Lysis buffer:
50ml
100ml
100mM NaCl(FW:58.44g)
.2922g
.5844g
10mM Tris-HCL pH7.5(FW:121.14g) or
1M Tris-HCL pH7.4
.061g
.5ml
.121g
1ml
50mM EDTA or
.5M EDTA pH 8.0
5ml
10ml
1% SDS
.5g
1g