AlanFung:LabNotes/Capturing/2012-6-22

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  • Perform one step qPCR using Indexing seq. primer
Content Volume MM
Captured DNA 10 Do not add to MM
2X Kapa SYBG qPCR master mix 50 110
10uM AmpF6.3Sol 4 8.8
10uM AmpR6.3Ind 4 Do not add to MM
H2O 32 70.4
   98C 30S -> (98C 10S -> 58C 20S -> 72C 20S) x 6 -> (98C 10S -> 72C 20S) x40
   Monitoring the reactions and terminate the program right before the amplification curves reach the plateau.
Ind1-H7gDNA
Ind2-H14gDNA

File:ZhangLab 2 2012-06-23 18hr 06min 1-6.jpg

  • Calculate amplicon size
AmpF6.3Sol AATGATACGGCGACCACCGACACTCTCAGATGTTATCGAGGTCCGAC
AmpR6.3Ind1  CAAGCAGAAGACGGCATACGAGATCGTGATGCTAGGAACGATGAGCCTCCAAC