Daniel:Notebook/HiResChrPaint/2014-9-10

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Padlock Probes II[edit]

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Nanodrop Results[edit]

Sample 1:1 ng/uL 5:1 ng/uL 5:1 ng/uL Projected 10:1 ng/uL 10:1 ng/uL Projected Average ng/uL Standard Deviation ug in 100 uL
PPRL-A 1944.1 373 1865 184.4 1844 1884.4 52.8 188.4
PPRL-B 2064.8 421.7 2108.5 203 2030 2067.8 39.3 206.8

Usually after ethanol precipitation I get really high results, so I'll just say these are fine and continue as though I have a reasonable amount of material.

DpnII/USER Digestion[edit]

Reaction Table

Reagent PPRL-A1 (uL) PPRL-A2 (uL) PPRL-B1 (uL) PPRL-B2 (uL)
amplicon 50 50 50 50
ug Input 94.2 94.2 103.4 103.4
USER 5 5 5 5
nf H2O 30 30 30 30
Total 85 85 85 85
  1. Set up reaction according to table in a 0.2 mL PCR tube
  2. Incubate for 2 hours at 37C
  3. Add 10 uL DpnII buffer (10X)
  4. Incubate at 94C for 2 minutes
  5. Cool to 37C and incubate for 3 minutes
  6. Add 5 uL DpnII and incubate at 37C for 2 hours
  7. Heat inactivate DpnII at 65C for 20 minutes
  8. [[Daniel:Protocols/

Nanodrop Results[edit]

Sample PPRL-A1 PPRL-A2 PPRL-B1 PPRL-B2
ng/uL 154.1 131 153 163.7
ug in 40 uL 6.2 5.2 6.1 6.5
ug total       24.1

TBE Gel[edit]

  1. For each sample, 5 uL TBE, 2 uL 6x dye, 1 uL sample; Load 7.5 uL each
  2. Run gel at 220V for 25 minutes


Lane 1 2 3 4 5 6 7 8 9 10 11 12
Sample Low Mass Ladder PPRL-A, pre-digest 10:1 PPRL-A, pre-digest 40:1 PPRL-A1 PPRL-A2 PPRL-B, pre-digest 10:1 PPRL-B, pre-digest 40:1 PPRL-B1 PPRL-B2

Gel Image

File:2014-09-09-PPRL.jpg