Daniel:Notebook/ComboLock/2017-1-7: Difference between revisions

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>Djacobse
(Created page with "=Version 2 Oligos (Started Yesterday)= Back to Calendar ==Protocol== <ol start="8"> <li>Exonuclease Dig...")
 
>Djacobse
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<li>Add 15 uL master mix to each tube</li>
<li>Add 15 uL master mix to each tube</li>
<li>Add sample to new reaction tubes according to following table</li>
<li>Add sample to new reaction tubes according to following table</li>
{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext>
|- style="background-color:#92CDDC;font-size:12pt;font-weight:bold" align="center"
| width="170" height="30" | Reagent
| width="65" | uL Added
| width="65" | Master Mix (8.5X)
|- style="font-size:12pt"
| height="15"  valign="bottom" | Template
| align="center" align="center" valign="bottom" | 5
| align="center" valign="bottom" | NA
|- style="background-color:#BFBFBF;font-size:12pt"
| height="15"  valign="bottom" | RCA Primer (100 uM)
| align="center" align="center" valign="bottom" | 2.5
| align="center" align="center" valign="bottom" | 21.25
|- style="font-size:12pt"
| height="15"  valign="bottom" | dNTP (1 mM)
| align="center" align="center" valign="bottom" | 0.8
| align="center" align="center" valign="bottom" | 6.8
|- style="background-color:#BFBFBF;font-size:12pt"
| height="15"  valign="bottom" | 10X Buffer
| align="center" align="center" valign="bottom" | 2
| align="center" align="center" valign="bottom" | 17
|- style="font-size:12pt"
| height="15"  valign="bottom" | Phi29
| align="center" align="center" valign="bottom" | 1
| align="center" align="center" valign="bottom" | 8.5
|- style="background-color:#BFBFBF;font-size:12pt"
| height="15"  valign="bottom" | nfH2O
| align="center" align="center" valign="bottom" | 8.7
| align="center" align="center" valign="bottom" | 73.95
|- style="font-size:12pt"
| height="15"  valign="bottom" | Total
|style="font-weight:bold" align="center" align="center" valign="bottom" | 20
|style="font-weight:bold" align="center" align="center" valign="bottom" | 127.5
|}

Revision as of 19:27, 6 January 2017

Version 2 Oligos (Started Yesterday)

Back to Calendar

Protocol

  1. Exonuclease Digestion
    1. Mix 12.5 uL Exo I (20U/uL) and 2.5 uL ExoIII (100U/uL)
    2. Add 2 uL to each sample, mix by swirling pipette tip
    3. Incubate for 1 hour at 37C
    4. Heat kill by incubating for 5 min at 95C
  2. Rolling Circle Amplification
    1. Prepare 7X Master mix according to table below
    2. Add 15 uL master mix to each tube
    3. Add sample to new reaction tubes according to following table
    4. Reagent uL Added Master Mix (8.5X)
      Template 5 NA
      RCA Primer (100 uM) 2.5 21.25
      dNTP (1 mM) 0.8 6.8
      10X Buffer 2 17
      Phi29 1 8.5
      nfH2O 8.7 73.95
      Total 20 127.5