AlanFung:LabNotes/CTCF/2011-6-6

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Revision as of 20:38, 6 June 2011 by >Alan6017518 (→‎Setup PCR Reaction)
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Perform bisulfite conversion on 200ng of gDNA

Protocol

Setup PCR Reaction

  • Add the components on ice in order while mixing gently.
Component Amount per reaction x6.6
Distilled water 31 204.6
10X pfuTurbo Cx reaction buffer 5 33
dNTPs (10mM each DNTP) 1 6.6
DNA template/STD 5
CHR8 F Primer (3.3uM) 3
CHR8 R Primer (3.3uM) 3
SYBR Green (50X) 1 6.6
PfuTurbo Cx Hotstart DNA Polymerase (2.5U/ul) 1 6.6
Total reaction volume 50 257.4
  • Add 39ul master mix to primers and templates
    • Perform Manufacturer protocol for PfuTurbo Cx Hotstart DNA Polymerase
95C 2m
95C 30s
52C 30s
72C 2m
Plate Read
Go to step 2 repeat for 29 times
72C 10m
15C hold

Results