Chris:LabNotes/FateMapping/Calendar/2014/2014-10-16

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Padlock Probe Production (designed 9/22)[edit]

Background[edit]

Custom Array
CA-1127
90k_oligos_30Sept2014
69.89ng/ul 80ul -> 5.59ug
10/14/14

Procedure[edit]

  • I used the same protocol that Noi's been using for probe capture

Expansion PCR[edit]

  • Considering the length of the probes are 127bp, I calculated that the concentration in the tube is 1667.62nM
  • Mastermix recipe (want to run 2 rxn's)
Component             1x (ul)
Oligo Mix             5.997
KAPA FAST MM (2x)     50
100uM AP1V61U         0.40
100uM AP2V6           0.40
H2O                   43.203
  • Noi noticed that with her probe set, there was an undesirable strong smear below 100bp. I ran a gel (as shown below) in order to verify the expansion PCR worked:

File:Cw ZhangLab 2 2014-10-16 17hr 58min pp production exp pcr.jpg

  • Unfortunately, I also see a strong smear below 100bp, when I would expect a strong band at 127bp