Kun:LabNotes/ASE/2008-1-6
Jump to navigation
Jump to search
Exp.1 Real-time PCR on the capturing reactions from 01/02/08 and 01/03/08's reactions[edit]
x 20 x 2 Template: 10ul or 15ul 10X buffer: 9ul or 14ul 25mM dNTP: 0.8ul or 1.2ul 100uM AmpV6.2SolF 0.4ul or 0.6ul 100uM AmpV6.2SolR 0.4ul or 0.6ul 50X SYBGI: 0.4ul or 0.6ul JumpStart Taq 2ul or 3ul H2O: 77ul or 115ul
94C 3min -> (94C 45 sec -> 57C 1min -> 72C 1min) x 10 -> (94C 30min -> 72C 1min) x 8 -> 72C 3min -> 15C hold.
The amplification curves didn't come up at the end of PCR. Then I realized that I should be using the AmpSolF2/AmpSolR primer pair. I added 0.5ul of 100uM primers to each reaction, repeat the PCR. The amplification curves came up, but I saw many non-specific bands in the PAGE gel.
File:2008-01-07-eSNP84k-capture-PCR.png