Noi/NOTES/2014-8-21

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Revision as of 21:11, 21 August 2014 by >Noi (Created page with "* '''Link to calendar''' * 2014-08-13: Received MOMOD V4 Oligo pool from CustomAraay === Oligo info. === * Name: 90K_ologos_04Aug2014 (8-20-14) * Con...")
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Oligo info.

  • Name: 90K_ologos_04Aug2014 (8-20-14)
  • Conc. 153.49ng/ul,
  • Volume 80ul in TE buffer. Total amount 12.28ug
  • Info from Dr. Zhang's note
           Description               # oligos   Length      Amp. Primers          Adaptor Removal
  Set A:   BSPP LMS cluster probes    73,574     148nt          V6                   USER/DpnII
  Set B:   Selector LMS cluster       12,994     148nt          V4                   USER/DpnII
  Set C:   Selector chr21 unique       2,539     148nt      eMIP_CA_F/R          Nt.AlwI/Nb.BrsDI
  Total:                              89,107
  • I will be working on 73K probe set
  • Length 148 mer (MW = 148 * 330Da/bp = 48,840 Da or g/mole
  • Conc. 20.53ng/ul = 3.1427uM or 3142.7nM (153.49ng/ul/ 48,840g/mole)

Expansion PCR (Quick test)

  • I did a quick test of expansion PCR on the three subset.
  • I diluted 1st amplicon of each primer set (amplified by V4, V6, or eMIP_CA) into 2nM and will be as positive control for amplification.
    • V4 set: I used MONOD GP1V4 as template. Mix 10ul of 20nM 1st round amplicon MONOD GP1V4 with 90ul H2O
    • V6 set: I used MONOD V3 as template. Prepare 2nM template in the same way as V4 set.
    • eMIP_CA: I used oligo from LC sciences used in Nature methods 2012. Mix 20ul of 10nM 1st round amplicon 4K LC Sciences with 80ul H2O

95C 30sec -> (95C 30sec -> 54C 45sec-> 72C 45sec) X 18-> 72C 3min -> 15C hold