Kun:LabNotes/ASE/2007-10-18
Jump to navigation
Jump to search
Exp. 1 Perform PCR on CES22k and eSNP84k libraries to make MIP probes.[edit]
Reaction CES22k(100nM):
x 8 Template 0.2ul 1.6ul 10X buffer 10ul 80ul 10mM dNTP 2ul 16ul 100uM pAP1V6U 0.4ul 3.2ul 100uM phosAp2V6 0.4ul 3.2ul JumpStart Taq 2ul 16ul H2O 85ul 680ul
Reactions eSNP84k set1-5 (100nM):
x 8 x 5 Template 0.2ul 1.6ul x 5 10X buffer 10ul 80ul x 5 10mM dNTP 2ul 16ul x 5 100uM AP1V41U 0.4ul 3.2ul x 5 100uM Ap2V4 0.4ul 3.2ul x 5 JumpStart Taq 2ul 16ul x 5 H2O 85ul 680ul x 5
94C3min -> 20 cycles of (94C 45sec -> 58C 2min -> 72C 1min) -> 72C 3min -> 4C hold
File:2007-10-19-AgilentLib-PCR.png
Estimated yield: CES22k: 12.5ng/ul eSNP84k1: 11.5ng/ul eSNP84k2: 13ng/ul eSNP84k3: 8.5ng/ul eSNP84k4: 12.75ng/ul eSNP84k5: 15.25ng/ul
Ethanol Precipitation, 400ul reactions per tube, two tubes per sample. Resuspended 50ul dH2O, measured concentration with Nanodrop.
NaOAC, pH=5 NaOAC, pH=7 CES22k: 215ng/ul 246ng/ul eSNP84k1: 216ng/ul 184ng/ul eSNP84k2: 184ng/ul 289ng/ul eSNP84k3: 157ng/ul 290ng/ul eSNP84k4: 142ng/ul 256ng/ul eSNP84k5: 163ng/ul 221ng/ul