Text file started Oct 10, 1998

A very simple method to obtain DNA for a few PCRs

Bruce Herron and Lorraine Flaherty

The original protocol came from (I think) a BioTechniques paper based on typing people for cystic fibrosis mutations with blood spots. Our in-house protocol is quite different from this:

  1. Start by taking tail biopsies as you normally would, certainly 2–3 mm is suited to this purpose. Freeze the tails in Eppendorfs as you may want to go back to them as a source of more DNA if your goal is the identification of recombinants.

  2. Before stopping the bleeding of the tail wound spot blood onto grade 1 Whatman paper. For this purpose we have 10 X 25 cm strips: on the left side we put the mouse ID# (in pencil) on the right edge four spots of blood. You can put them about every two centimeters. Placing spots much closer will result in samples bleeding into each other.

  3. After the blood is dried, soak the sheet in 100% methanol for 5 minutes, then dry.

    Fixed DNA samples are now ready for PCR. To amplify punch out 2 mm holes (an ear punch works well) into PCR tubes. Add 10 µl water and heat for 5 minutes at 95 deg C, add PCR components and amplify. Others in the lab have eliminated the preheat step.

Two words of caution with this protocol. First, as with ear punch methods, there is cross contamination if you choose not to clean the punch between samples so this method can be a pain for typing in presence vs absence screens. Second, we found this method successful with radiolabled PCR assays, but the sensitivity with agarose typings was lower, but still functional with most primer pairs.

Bruce Herron
Developmental Genetics
David Axelrod Institute
120 New Scotland Ave.
Albany, NY 12208
FAX (518) 474-3181