Daniel:Notebook/ComboLock/2017-1-26: Difference between revisions

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>Djacobse
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<li>Add 47 uL master mix to each well</li>
<li>Add 47 uL master mix to each well</li>
<li>Add 2 uL sample and 1 uL appropriate Reverse Primer (1 uM) according to table</li>
<li>Add 2 uL sample and 1 uL appropriate Reverse Primer (1 uM) according to 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="100" height="45" | Sample
| width="100" | Condition
| width="65" | Lane
| width="65" | AmpR Index
| width="65" | Sample Vol (uL)
| width="85" | 2X Kapa SYBR qPCR MM
| width="65" | 10 uM Forward Primer
| width="65" | 10 uM Reverse Primer
| width="65" | H2O
| width="65" | Total Volume (uL)
|- style="font-size:12pt"
|style="font-weight:bold" height="15"  | Sample 6AC
| align="center" | Epicentre
| align="center" | A1
| align="center" align="center" | 22
| align="center" align="center" | 2
| align="center" align="center" | 25
| align="center" align="center" | 1
| align="center" align="center" | 1
| align="center" align="center" | 21
| align="center" align="center" | 50
|- style="background-color:#BFBFBF;font-size:12pt"
|style="font-weight:bold" height="15"  | Sample 6BC
| align="center" | Epicentre
| align="center" | A2
| align="center" align="center" | 22
| align="center" align="center" | 2
| align="center" align="center" | 25
| align="center" align="center" | 1
| align="center" align="center" | 1
| align="center" align="center" | 21
| align="center" align="center" | 50
|- style="font-size:12pt"
|style="font-weight:bold" height="15"  | Sample 6AD
| align="center" | Epicentre 1:10
| align="center" | A3
| align="center" align="center" | 23
| align="center" align="center" | 2
| align="center" align="center" | 25
| align="center" align="center" | 1
| align="center" align="center" | 1
| align="center" align="center" | 21
| align="center" align="center" | 50
|- style="background-color:#BFBFBF;font-size:12pt"
|style="font-weight:bold" height="15"  valign="bottom" | Sample 6BD
| align="center" | Epicentre 1:10
| align="center" | A4
| align="center" align="center" | 23
| align="center" align="center" | 2
| align="center" align="center" | 25
| align="center" align="center" | 1
| align="center" align="center" | 1
| align="center" align="center" | 21
| align="center" align="center" | 50
|- style="font-size:12pt"
|style="font-weight:bold" height="15"  valign="bottom" | NTC
| align="center"  valign="bottom" | &nbsp;
| align="center" valign="bottom" | A5
| align="center" align="center" valign="bottom" | 24
| align="center" align="center" valign="bottom" | 2
| align="center" align="center" valign="bottom" | 25
| align="center" align="center" valign="bottom" | 1
| align="center" align="center" valign="bottom" | 1
| align="center" align="center" valign="bottom" | 21
| align="center" align="center" valign="bottom" | 50
|}


<li>qPCR Cycles</li>
<li>qPCR Cycles</li>

Revision as of 21:10, 26 January 2017

RCA Test (Started Monday)

Back to Calendar

Epicentre Test (Started Yesterday)

Sample Matrix

  1. qPCR
    1. Make 5.2X qPCR master mix according to following recipe
      1. 109.2 uL nfH2o
      2. 5.2 uL AmpF</li
      3. 130 uL 2X Kapa SYBR Fast Master Mix
      4. Add 47 uL master mix to each well
      5. Add 2 uL sample and 1 uL appropriate Reverse Primer (1 uM) according to table
      6. Sample Condition Lane AmpR Index Sample Vol (uL) 2X Kapa SYBR qPCR MM 10 uM Forward Primer 10 uM Reverse Primer H2O Total Volume (uL)
        Sample 6AC Epicentre A1 22 2 25 1 1 21 50
        Sample 6BC Epicentre A2 22 2 25 1 1 21 50
        Sample 6AD Epicentre 1:10 A3 23 2 25 1 1 21 50
        Sample 6BD Epicentre 1:10 A4 23 2 25 1 1 21 50
        NTC   A5 24 2 25 1 1 21 50
      7. qPCR Cycles
        1. 95C 3 min
        2. 95C 3 sec
        3. 55C 30 sec
        4. 72C 20 sec
        5. plate read
        6. goto b x24
        7. 72C 2 min
        8. 16C hold
    2. TBE Gel
      1. Mix 64 uL TBE, 16 uL 6x loading dye
      2. Aliquot 10 uL per sample/ladder lane onto parafilm
      3. Add 2 uL of sample or ladder to correct drop
      4. Load 10 uL in to well
      5. Run gel for 23 minutes at 230V
      6. Open gel and stain with 3 uL SYBR Gold for 3 minutes
      7. Rinse gel and image in gel doc

    Results