PCR

For a 25 ul reaction add:

  1. H2O 10.5 ul
  2. Taq 2X Master Mix 12.5 ul
  3. Forward primer 0.5 ul 100 uM solution
  4. Reverse primer 0.5 ul 100 uM solution
  5. DNA template 1 ul

General PCR cycling conditions are:

  1. An initial 95C 30s
  2. Then 30 cycles of:
    1. 95C 20s melting
    2. annealing temperature  (this varies with the primers used, usually between 55C and 65C) 40s
    3. 70C 1m extension (this can be raised to 5 m for several kbp long segments)
  3. With a final 70C of 5 minutes.

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