Reliable noninvasive genotyping based on excremental PCR of nuclear DNA purified with a magnetic bead protocol.
A new protocol for extraction of DNA from faeces is presented. The protocol involves gentle washing of the surface of the faeces followed by a very simple DNA extraction utilizing the wash supernatant as the source of DNA. Unlike most other protocols, it does not involve the use of proteinase K and/or organic extraction, but is instead based on adsorption of the DNA to magnetic beads. The protocol was tested by microsatellite genotyping across six loci for sheep and reindeer faeces. Comparison with DNA extracted from blood demonstrated that the protocol was very reliable, even when used on material stored for a long time. The protocol was compared with another simple, solid-phase DNA-binding protocol, with the result that the bead-based protocol gave a slightly better amplification success and a lower frequency of allelic drop-outs. Furthermore, our experiments showed that the surface wash prior to DNA extraction is a crucial step, not only for our protocol, but for other solid-phase protocols as well.
Flagstad
O
O
Department of Biology, University of Oslo, Norway.
Røed
K
K
Stacy
J E
JE
Jakobsen
K S
KS
D000818:Animals; D004247:DNA / Q000302:isolation & purification*; D005243:Feces / Q000737:chemistry*; D005838:Genotype; D008280:Magnetics; D018895:Microsatellite Repeats / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012053:Reindeer / Q000145:classification / Q000235:genetics; D012418:Ruminants / Q000145:classification* / Q000235:genetics; D012756:Sheep / Q000145:classification / Q000235:genetics
D004247:DNA