Daniel:Notebook/HiResChrPaint/2014-7-8: Difference between revisions
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>Djacobse (Created page with "=Strand Control= Back to Calendar Andrew has pointed out to me that I should always do the processing on the probes, just to keep results c...") |
>Djacobse No edit summary |
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I'll be running through V6S2 and V6S3, my favorites. | I'll be running through V6S2 and V6S3, my favorites. | ||
#Reaction Table | |||
{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext> | |||
|- style="font-size:12pt" align="center" | |||
|style="background-color:#FFFF00" align="center" width="81" height="45" | | |||
|style="font-weight:bold" width="65" | 10 nM OligoPool | |||
|style="font-weight:bold" width="65" | 2X Kapa SYBR qPCR MM | |||
|style="font-weight:bold" width="65" | 100 uM AP1V61U | |||
|style="font-weight:bold" width="65" | 100 uM AP2V6 | |||
|style="font-weight:bold" width="65" | H20 | |||
|style="font-weight:bold" width="65" | Total Volume | |||
|- style="background-color:#8DB4E2;font-size:12pt" align="center" | |||
| height="15" | per rxn | |||
| align="center" | 0.4 | |||
| align="center" | 50 | |||
| align="center" | 0.4 | |||
| align="center" | 0.4 | |||
| align="center" | 48.8 | |||
| align="center" | 100 | |||
|- style="font-size:12pt" align="center" | |||
|style="font-weight:bold" height="15" | V6S2 (12.2x) | |||
| align="center" | 4.88 | |||
| align="center" | 610 | |||
| align="center" | 4.88 | |||
| align="center" | 4.88 | |||
| align="center" | 595.36 | |||
| align="center" | 1220 | |||
|- style="font-size:12pt" align="center" | |||
|style="font-weight:bold" height="15" | V6S3 (12.2x) | |||
| align="center" | 4.88 | |||
| align="center" | 610 | |||
| align="center" | 4.88 | |||
| align="center" | 4.88 | |||
| align="center" | 595.36 | |||
| align="center" | 1220 | |||
|} | |||
#Aliquot 100 uL into each of 12 qPCR tubes | |||
#PCR protocol: | |||
##PCR 3 min 95C | |||
##95C 3 sec | |||
##55C 30 sec | |||
##72C 20 sec | |||
##plate read | |||
##goto ii x17 | |||
##72C 2 min | |||
##16C hold | |||
#Pool 3 samples together | |||
#Column Purification, elute with 40 uL EB | |||
#Measure concentrations in nanodrop |
Revision as of 22:31, 8 July 2014
Strand Control
Andrew has pointed out to me that I should always do the processing on the probes, just to keep results consistent. I am fairly confident this is not the cause of my difficulties, but it is still wise to keep that particular aspect consistent. As such, I will be running through the more rigorous method of probe prep.
qPCR
I'll be running through V6S2 and V6S3, my favorites.
- Reaction Table
10 nM OligoPool | 2X Kapa SYBR qPCR MM | 100 uM AP1V61U | 100 uM AP2V6 | H20 | Total Volume | |
per rxn | 0.4 | 50 | 0.4 | 0.4 | 48.8 | 100 |
V6S2 (12.2x) | 4.88 | 610 | 4.88 | 4.88 | 595.36 | 1220 |
V6S3 (12.2x) | 4.88 | 610 | 4.88 | 4.88 | 595.36 | 1220 |
- Aliquot 100 uL into each of 12 qPCR tubes
- PCR protocol:
- PCR 3 min 95C
- 95C 3 sec
- 55C 30 sec
- 72C 20 sec
- plate read
- goto ii x17
- 72C 2 min
- 16C hold
- Pool 3 samples together
- Column Purification, elute with 40 uL EB
- Measure concentrations in nanodrop