WangLab:PCR Programs

From OpenWetWare

Revision as of 16:29, 23 May 2007 by Lu6007 (Talk | contribs)
(diff) ←Older revision | Current revision (diff) | Newer revision→ (diff)
Jump to: navigation, search

Contents

Program 5 (single mutation for quickchange, 5-6kbp, Pfu polymerase, for PCDNA3 or PCDNA3’)

  1. stage 1, step 1, 95oC, 30 sec, cycle 1
  2. stage 2, for 16 cycles.
    • step 1, 95oC, 30 sec
    • step 2, 55oC, 1 min
    • step 3, 68oC, 18 min

Program 4 (single mutation for quickchange, 5-6kbp, Pfu polymerase, for PRSETb)

  1. stage 1, step 1, 95oC, 30 sec, cycle 1
  2. stage 2, for 18 cycles.
    • step 1, 95oC, 30 sec
    • step 2, 55oC, 1 min
    • step 3, 68oC, 12 min

Program 32 (PCR SH2 domain, 5-6kbp)

  1. stage 1, for 2 cycles, hold temperature.
    • step 1, 95oC, 2 min,
    • step 2, 40oC, 2 min
    • step 3, 72oC, 2 min 30 sec
  2. stage 2, for 28 cycles
    • step 1, 95oC, 1 min
    • step 2, 52oC, 1 min
    • step 3, 72oC, 2 min 30 sec
  3. stage 3, step 1, 72oC, 10 min

Program 10 (PCR for difficult annealing primers)

  1. stage 1, step 1, 95oC, 5 min
  2. stage 2, cycles 2.
    • step 1, 95oC, 1 min
    • step 2, 40oC, 1 min
    • step 3, 72oC, 2 min 30 sec
  3. stage 3, cycles 5
    • step 1, 95oC, 1 min
    • step 2, 43oC, 1 min
    • step 3, 72oC, 2 min 30 sec
  4. stage 4, cycles 5
    • step 1, 95oC, 1 min
    • step 2, 45oC, 1 min
    • step 3, 72oC, 2 min 30 sec
  5. stage 5, cycles 20
    • step 1, 95oC, 1 min
    • step 2, 52oC, 1 min
    • step 3, 72oC, 2 min 30 sec
  6. stage 6, step 1, 72oC, 10 min, cycle 1
Personal tools