Noi/NOTES/2013-2-27

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Plan for probe preparation for Matt's project[edit]

  • I will be preparing probes using the same method as I did for LC Sciences oligoes, [[1]]
  • Check reagents to make sure I have all reagents ready for probe prep and padlock capture
Probe prep
- Waiting for seed oligos: expected to come next Wednesday (March6, 2013)
- eMIP_CA1_F and eMIP_CA1_R primer for expansion PCR and production PCR (very little left, ordered 2/28/2013) 
  ; arrived 3/04/2013 -> resuspend 100uM
- 2x KAPA SYBG FAST qPCR MM
- Nt.AlwI (NEB) and Nb.BsrDI (NEB) -> ordered 2/26/2013 ; arrived 3/04/2013
- Qiaquick columns, TBU gel, Nanosep columns
Padlock capture
Note: There is no gap for this capture, so only need Ampligase
- Ampligase and buffer: yes
- ExoI/ExoIII: yes
- Amplification primers (Matt)
  • Look at the probe design (primer region), it's pretty much the same as the probes synthesized by LC Sciences oligoes. Start at AGGACCGGATCAACT
  • Need to check with Dr. Zhang about the gDNA, RNA and cDNA amount in capture. The expected size of citcularized DNA (no gap --> 170bp plus length of Illumina adapter = XX bp? look at primer design)
  • Continued on: http://genome-tech.ucsd.edu/LabNotes/index.php/Noi/NOTES/2013-3-6