AlanFung:LabNotes/Sequencing/2009-12-5

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qPCR Quantification

PGP4 PGP6 PGP8 PGP9 PGP8D PGP8E Dpn-CV-iF2 Dpn-CV-F2

Setup
Amp Phi-X (1nM) PGP6 PGP9 PGP8E Dpn-CV-iF2 (1/10)
Amp Phi-X (1nM) PGP6 PGP9 PGP8E Dpn-CV-iF2 (1/10)
Amp Phi-X (1nM) PGP6 PGP9 PGP8E Dpn-CV-iF2 (1/10)
Amp Phi-X (1nM) PGP6 PGP9 PGP8E Dpn-CV-iF2 (1/10)
PGP4 PGP8 PGP8D Dpn-CV-F2 (1/10)
PGP4 PGP8 PGP8D Dpn-CV-F2 (1/10)
PGP4 PGP8 PGP8D Dpn-CV-F2 (1/10)
PGP4 PGP8 PGP8D Dpn-CV-F2 (1/10)
x18 *1.1
DNA 2
Syb_FP5 (100uM) 0.2 3.6 3.96
Syb_RP7 (100uM) 0.2 3.6 3.96
2X Phusion 25 450 495
50X SYBR Green I 0.2 3.6 3.96
H20 24 432 475.2
Total 982.08
  • Dilute sample, 1ul in 9ul ddh2o
  • mix 2ul 1/10 sample with 98ul mix
  • aliquot 24ul to each well
98C 30 sec -> 20 cycles of (98C 10sec->64C 20sec->72C 20sec)->72C 5 min
Average Difference 2^ nM
Amp Phi-X (1nM) 9.833 9.899 9.819 9.990 9.89 0
PGP4 9.471 9.522 9.562 9.706 9.57 0.32 1.25 1.25
PGP6 10.667 10.911 11.011 10.86 0.97 1.96 0.51
PGP8 9.719 9.813 10.088 10.166 9.95 0.06 1.04 0.96
PGP9 9.663 9.710 9.751 9.71 0.18 1.13 1.13
PGP8D 5.250 5.262 5.287 5.27 4.62 24.65 24.65
PGP8E 5.067 5.143 5.168 5.13 4.76 27.17 27.17
Dpn-CV-F2 (1/10) 8.303 8.403 8.663 8.666 8.51 1.38 2.6 26
Dpn-CV-iF2 (1/10) 9.824 9.962 10.000 9.93 0.04 1.03 9.74