Amplification of Microsatellites Resolution and Detection

The details given in this section assume that the fluorescently labeled primer method will be followed. See Note 2 for details for the radioactive and fluorescent dCTP incorporation methods.

1. Genomic DNA from tumor and normal tissue.

2. dNTP mix (Pharmacia).

3. 10X PCR buffer (supplied with enzyme).

4. MgCl2 solution (supped with enzyme).

5. Thermoprime Taq DNA polymerase (Applied Biotechnologies, UK).

6. Microsatellite primers (see Note 3; Oswel, UK).

7. 0.2-mL Microcentrifuge tubes (Perkin Elmer).

8. 0.5-mL Genetic Analyser sample tubes (Applied Biosystems).

9. Septa for 0.5-mL Sample tubes (Applied Biosystems). 10. PCR machine (Perkin Elmer).

II. ABI PRISM 310® Genetic Analyser (Applied Biosystems; see Note 4).

12. De-ionised formamide (Applied Biosystems).

13. TAMRA-500 size standard (Applied Biosystems).

14. GeneScan® Analysis Software V3.1 (Applied Biosystems; see Note 4).

Was this article helpful?

0 0

Post a comment