Kun:LabNotes/CpgSeq/2010-6-8

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Probe prep for the DMR220k set

          Library name       OLID     Part #      
            DMR220k_a       0284601   G4446A    
            DMR220k_b       0284591   G4446A    
            DMR220k_c       0284581   G4446A    
            DMR220k_d       0284571   G4446A   
  • Resuspend the oligos in each tube with 500ul nuclease-free H2O.

Expansion PCR

                               x 4
 DMR220k_a     (20nM)          1ul                               
 DMR220k_b     (20nM)          1ul
 DMR220k_c     (20nM)          1ul                               
 DMR220k_d     (20nM)          1ul
 2x iTaq master mix:          100ul  
 100uM AP1V6:                 0.8ul
 100uM AP2V6:                 0.8ul
 H2O:                          96ul
 94C 2min -> (94C 30sec -> 54C 45sec -> 72C 45sec) x 16 -> 72C 3min -> 15C hold  
 Purify the amplicons with two Qiaquick columns, eluted with 40ul EB.

Production PCR

Digestion and purification