Daniel:Notebook/PosSequencing/2017-6-13: Difference between revisions

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>Djacobse
>Djacobse
Line 21: Line 21:
<li>870 (60) uL 2X Kapa SYBR</li>
<li>870 (60) uL 2X Kapa SYBR</li>
<li>638 (44) uL nfH<sub>2</sub>O</li></ol>
<li>638 (44) uL nfH<sub>2</sub>O</li></ol>
<li>Add 28 uL master mix according to plate (4X master mix is for positive controls, 58X is for samples and negative controls)</li>
<li>Add 28 uL master mix according to plate layout (4X master mix is for positive controls, 58X is for samples and negative controls)</li>
 
[[Image:PlateLayout-20170613-BiotinTest.png|350x250px]]
 
<li>Add 2 uL appropriate sample according to plate layout</li>
<li>Add 2 uL appropriate sample according to plate layout</li>
<li>Use "standard" thermocycling program</li>
<li>Use "standard" thermocycling program</li>
</ol>
</ol>

Revision as of 18:48, 13 June 2017

Biotin vs Dual Biotin (Started Yesterday)

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Workflow Reminder

File:ExperimentMap-20170610-Biotin-DualBiotin.png

Protocol

  1. qPCR
    1. Prepare 2 plates; 1 single biotin and 1 dual biotin
    2. For each sample prepare 58X master mix and (4X) master mix: 1X components set to 30 uL; add 2uL sample
      1. 58 (4) uL 1 uM primer2
      2. (4) uL 1 uM biotin/2biotin uracil primer
      3. 58 uL 10 pM barcode oligo v2
      4. 870 (60) uL 2X Kapa SYBR
      5. 638 (44) uL nfH2O
    3. Add 28 uL master mix according to plate layout (4X master mix is for positive controls, 58X is for samples and negative controls)
    4. File:PlateLayout-20170613-BiotinTest.png
    5. Add 2 uL appropriate sample according to plate layout
    6. Use "standard" thermocycling program