Rui:LabNotes/Collaborations/2011-5-20: Difference between revisions
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==Additional 10 cycles of direct lysis samples== | ==Additional 10 cycles of direct lysis samples== | ||
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===QPCR=== | ===QPCR=== | ||
[[File:5.20.11_remaining samples.jpg]] | [[File:5.20.11_remaining samples.jpg]] | ||
===Beads purification=== | |||
* Combined into 4 samples, 8 PCR tubes per samples | |||
* Sperm RNA, ES RNA, Sperm lysis, ES lysis | |||
* 0.7X beads purification | |||
* Elute with 50ul water | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Sample ID''' | |||
| align="center" style="background:#f0f0f0;"|'''ng/ul''' | |||
| align="center" style="background:#f0f0f0;"|'''260/280''' | |||
| align="center" style="background:#f0f0f0;"|'''260/230''' | |||
|- | |||
| sperm RNA||43.64||1.9||1.68 | |||
|- | |||
| ES RNA||295.67||1.93||2.18 | |||
|- | |||
| ES lysis||220.08||1.92||1.97 | |||
|- | |||
| sperm lysis||23.5||1.68||0.95 | |||
|- | |||
| | |||
|} | |||
===Cloning=== | |||
* Ligation: ~25-30ng purified PCR + 2.5ng vector in 10ul total ligation, 14C, o/n | |||
* Transformation, using Kan30 plate |
Latest revision as of 18:21, 17 June 2011
Additional 10 cycles of direct lysis samples[edit]
- Continue with lab note on 5.19.11
- Additional 10 cycles for direct lysis samples (sperm and ES) in parallel with 500pg RNA and 150pg gDNA
QPCR curve[edit]
File:Additional 10 cycles 5.20.11.jpg
TBE PAGE[edit]
Repeat 2nd strands and PCR with the remaining samples[edit]
- Continue with samples on 5.19.11, the remaining 9ul A-tailing product
- 2nd strand with P8
- PCR with 30 cycles
QPCR[edit]
File:5.20.11 remaining samples.jpg
Beads purification[edit]
- Combined into 4 samples, 8 PCR tubes per samples
- Sperm RNA, ES RNA, Sperm lysis, ES lysis
- 0.7X beads purification
- Elute with 50ul water
Sample ID | ng/ul | 260/280 | 260/230 |
sperm RNA | 43.64 | 1.9 | 1.68 |
ES RNA | 295.67 | 1.93 | 2.18 |
ES lysis | 220.08 | 1.92 | 1.97 |
sperm lysis | 23.5 | 1.68 | 0.95 |
Cloning[edit]
- Ligation: ~25-30ng purified PCR + 2.5ng vector in 10ul total ligation, 14C, o/n
- Transformation, using Kan30 plate