Biotech 2nd Edition Sample Ch. 4 Biotechnology Sample Ch. 4 - Page 10

Procedure EtBr Dot Test for DNA a. Clean and dry the glass surface of a transilluminator light box. b. Add 10 µL of sample to be tested to the transilluminator glass surface. c. Add 10 µL of EtBr solution to the sample. Pipet the mixture up and down once to mix. d. Other samples may be tested at other spots on the illuminator glass surface. Make sure the “dot” of sample is kept apart from other samples. e. Close the lid and turn on the UV lamp. f. Observe the amount of color and glowing of each sample compared with the positive (2 mg/mL of salmon sperm stock solution) and negative (deionized water) standards, each mixed with 10 µL of EtBr. 1. Using the EtBr dot-test procedures, test each (unknown) sample thought to contain DNA. Also, test deionized water as a negative control, and test 2 mg/mL of salmon sperm DNA solution as a positive control. 2. Photograph and evaluate the results. Use a 5 → 0 scale to assign a numerical rating to the relative amount of DNA in the sample compared with the controls (see Figure 4.5). Record the results in a data table in your notebook. Data Analysis/Conclusion Estimate the concentration of the unknown samples. Do this by converting the numerical rating they were given to an estimated concentration (mg/mL) compared wi H