Kun:LabNotes/CpgSeq/2007-12-11

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Exp.1 Repeat the PCR experiment on the circularized reactions using Hd48MIP1,2,5 primers with a touchdown protocol

   Template:            1ul  (1:100 dilution of bifulfite converted gDNAs)
   10X buffer:        2.5ul
   10mM dNTP:         0.5ul
   10uM Primer pair  0.75ul
   50X SYBR I         0.2ul
   AmpliTaq Gold     0.25ul
   H2O                 20ul

Total number of reactions: 4 MIP reactions x 3 primer pairs + 2 control x 3 primer pairs = 18 95C 15min -> 8 cycles of (94C 30sec -> 60C 30sec (-0.5C per cycle) -> 72C 30 sec) -> 27 cycles of (94C 30sec -> 56C 30sec -> 72C 30 sec) -> 72C 3min -> 15C hold.