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SCOC Multiplex 1
PCR
- multiplex looks good in some PCRs (when run next to uniplexes), but not in others (when scaled up for frag submission)
- try full gradient (temperature and buffer) of multiplexed reaction
Gel
| Pour |
Load |
Run
|
| 270 mL 1x TAE + 5.4 g agarose |
1 µL gel load dye + 5 µL PCR product |
160 V
|
| 27 µL GelRed |
6 µL ladder |
50 minutes
|
 - not favorable results
- lane for Buffer E, 64.3° consistent with 23 June 2011 PCR/gel
- in most lanes, can see the Scun9/10/11 band
- some lanes tend to have larger (Scun15, 22) or small amplicons (Scun2, 3)
- given that this doesn't seem to be able to get 'better', will submit yesterday's PCR to UAGC
- should be able to see which loci aren't amplifying, which loci are dominant
- might be able to split the multiplex into two separate multiplexes
- looking at Gradient Summary, 13 June 2011:
- Scun3, 9, 11 - Buffer D, 64°C
- Scun2, 10, 15, 22 - Buffer D, 56°C
- Scun10, 15, 22 - Buffer G, 61°C
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Tara's Lab Notebook
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