Protocol: Restriction digestion of Phytophthora infestans DNA for RG57 fingerprinting
- Determine the approximate concentration of your DNA sample by running a portion of the sample on a gel. We usually run 5 ul of a 50 ul DNA sample.
- Label a 1.5 ml microfuge tube for each digestion.
- Prepare a digestion mixture for each sample in the labelled 1.5 ml tube.
| 1-2 ug DNA | x ul |
| spermadine (10 mg/ml) | 2.0 ul |
| 10X enzyme buffer | 4.0 ul |
| Eco R1 enzyme | 2.0 ul |
| RNase A (10 mg/ml) | 1.0 ul |
| H2O | to 40 ul |
- Place the tubes in the 37° C incubator for at least 6 hr or overnight.
- Add running dye to each sample, load onto a 0.8 % agarose gel and run overnight at 30-40 volts.
