Kun:LabNotes/ASE/2007-11-2

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Exp. 1 Perform PCR on the eSNP84k set using different Tag polymerases.[edit]

  • ABI Platinium Taq Gold
  • Thermo Start Taq
  • Supernova HotStart
  • Qiagen HotStar Plus
  • Bio-Rad iTaq
  • Sigma JumpStart Taq

Template: eSNP84k set #5.

I use the same JumpStart 10X buffer.

        template:       0.2ul         1.6ul x 6
        10x buffer:      10ul          80ul x 6
        10mM dNTP:        2ul          16ul x 6
        100uMAP1V41U:   0.4ul         3.2ul x 6
        100uMphosAP2V4: 0.4ul         3.2ul x 6
        Taq:            0.5 or 1ul   8 or 16ul x 6 
        50x SYBR I:     0.8ul         6.4ul x 6
        H2O:             86ul         680ul x 6

95C 12min (or 5min) -> 94C 3min -> 17 cycles of (94C 45sec -> 58C 2min -> 72C 1min) -> 72C 3min -> 15C hold.

Surprisingly, all reactions looked good based on amplification curves. I added two more cycles since the curves have not reached to plateau yet.