Daniel:Notebook/ComboLock/2017-2-2: Difference between revisions
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>Djacobse (Created page with "=RCA Time Test (Started Friday Jan 27)= Back to Calendar ==Phosphorothiorate RCA Primer (Started [[Dani...") |
>Djacobse |
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<li>Add 2 uL sample and 1 uL AmpR indexed according to table</li> | <li>Add 2 uL sample and 1 uL AmpR indexed 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="89" height="45" | Sample | |||
| width="65" | Sample | |||
| width="77" | Condition | |||
| width="65" | AmpR Index | |||
| width="65" | Lane | |||
| width="65" | Sample Vol (uL) | |||
| width="65" | 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 1AN | |||
| align="center" | Sample | |||
| align="center" | 3 hours | |||
| align="center" align="center" | 20 | |||
| align="center" | A8 | |||
| 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 1BN | |||
| align="center" | Sample | |||
| align="center" | 3 hours | |||
| align="center" align="center" | 20 | |||
| align="center" | A7 | |||
| 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 3AN | |||
| align="center" | No C Probe | |||
| align="center" | 3 hours | |||
| align="center" align="center" | 21 | |||
| align="center" | A6 | |||
| 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 3BN | |||
| align="center" valign="bottom" | No C Probe | |||
| align="center" | 3 hours | |||
| align="center" align="center" | 21 | |||
| align="center" | A5 | |||
| 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 1AO | |||
| align="center" | Sample | |||
| align="center" | 3 hours-Epi | |||
| align="center" align="center" | 22 | |||
| align="center" | A4 | |||
| 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 1BO | |||
| align="center" | Sample | |||
| align="center" | 3 hours-Epi | |||
| align="center" align="center" | 22 | |||
| align="center" | A3 | |||
| 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 3AO | |||
| align="center" | No C Probe | |||
| align="center" | 3 hours-Epi | |||
| align="center" align="center" | 23 | |||
| align="center" | A2 | |||
| 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 3BO | |||
| align="center" valign="bottom" | No C Probe | |||
| align="center" | 3 hours-Epi | |||
| align="center" align="center" | 23 | |||
| align="center" | A1 | |||
| 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" | NTC | |||
| align="center" align="center" | | |||
| align="center" align="center" | | |||
| align="center" align="center" | 24 | |||
| align="center" | H1 | |||
| align="center" align="center" | 0 | |||
| align="center" align="center" | 25 | |||
| align="center" align="center" | 1 | |||
| align="center" align="center" | 1 | |||
| align="center" align="center" | 23 | |||
| align="center" align="center" | 50 | |||
|} | |||
<li>qPCR Cycles</li> | <li>qPCR Cycles</li> | ||
Line 31: | Line 163: | ||
<li>TBE Gel</li> | <li>TBE Gel</li> | ||
<ol type="A"> | <ol type="A"> | ||
<li>Mix | <li>Mix 94 uL TBE, 24 uL 6x loading dye</li> | ||
<li>Aliquot 10 uL per sample/ladder lane onto parafilm</li> | <li>Aliquot 10 uL per sample/ladder lane onto parafilm</li> | ||
<li>Add 2 uL of sample or ladder to correct drop</li> | <li>Add 2 uL of sample or ladder to correct drop</li> | ||
<li>Load 10 uL in to well</li> | <li>Load 10 uL in to well</li> | ||
<li>Run gel for | <li>Run gel for 23 minutes at 230V</li> | ||
<li>Open gel and stain with 3 uL SYBR Gold for 3 minutes</li> | <li>Open gel and stain with 3 uL SYBR Gold for 3 minutes</li> | ||
<li>Rinse gel and image in gel doc</li> | <li>Rinse gel and image in gel doc</li> |
Revision as of 17:47, 2 February 2017
RCA Time Test (Started Friday Jan 27)
Phosphorothiorate RCA Primer (Started Yesterday)
- qPCR
- Make qPCR master mix according to following recipe
- 361.2 uL nfH2O
- 17.2 uL AmpF6.4
- 430 uL Kapa SYBR Fast
- Add 47 uL master mix to each well
- Add 2 uL sample and 1 uL AmpR indexed according to table
- qPCR Cycles
- 95C 3 min
- 95C 3 sec
- 55C 30 sec
- 72C 20 sec
- plate read
- goto b x24
- 72C 2 min
- 16C hold
- TBE Gel
- Mix 94 uL TBE, 24 uL 6x loading dye
- Aliquot 10 uL per sample/ladder lane onto parafilm
- Add 2 uL of sample or ladder to correct drop
- Load 10 uL in to well
- Run gel for 23 minutes at 230V
- Open gel and stain with 3 uL SYBR Gold for 3 minutes
- Rinse gel and image in gel doc
Sample | Sample | Condition | AmpR Index | Lane | Sample Vol (uL) | 2X Kapa SYBR qPCR MM | 10 uM Forward Primer | 10 uM Reverse Primer | H2O | Total Volume (uL) |
Sample 1AN | Sample | 3 hours | 20 | A8 | 2 | 25 | 1 | 1 | 21 | 50 |
Sample 1BN | Sample | 3 hours | 20 | A7 | 2 | 25 | 1 | 1 | 21 | 50 |
Sample 3AN | No C Probe | 3 hours | 21 | A6 | 2 | 25 | 1 | 1 | 21 | 50 |
Sample 3BN | No C Probe | 3 hours | 21 | A5 | 2 | 25 | 1 | 1 | 21 | 50 |
Sample 1AO | Sample | 3 hours-Epi | 22 | A4 | 2 | 25 | 1 | 1 | 21 | 50 |
Sample 1BO | Sample | 3 hours-Epi | 22 | A3 | 2 | 25 | 1 | 1 | 21 | 50 |
Sample 3AO | No C Probe | 3 hours-Epi | 23 | A2 | 2 | 25 | 1 | 1 | 21 | 50 |
Sample 3BO | No C Probe | 3 hours-Epi | 23 | A1 | 2 | 25 | 1 | 1 | 21 | 50 |
NTC | 24 | H1 | 0 | 25 | 1 | 1 | 23 | 50 |