Matt:LabNotes/2015-3-25: Difference between revisions
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>Mzcai (Created page with "==CA12k_Nov2014_V4 Probe Production== *Production of Padlock Probes (V4) Matt:LabNotes/2014-11-19#Assembly_of_oligo_pool_for_ordering *Do double 96 100ul PCR Reactions ==...") |
>Mzcai mNo edit summary |
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'''Program''' 98C 1min -> (98C 10sec -> 55C 30sec -> 72C 45sec) x 15 -> 72C 2min -> 15C hold | '''Program''' 98C 1min -> (98C 10sec -> 55C 30sec -> 72C 45sec) x 15 -> 72C 2min -> 15C hold | ||
[[File: | 800px]] | [[File:032515_CA12kNov14_ProductionPCR_V4.JPG | 800px]] | ||
===Zymo DNA Clean & Concentrator 100=== | |||
*Purify 10ml of PCR product with 1 Zymo-spin V Column/Reservoir | |||
**Add 50ml DNA Binding Buffer to 10ml product (Use 2 50ml tubes) | |||
*Elute with 540ul H2O | |||
*Measured concentration with Nanodrop: | |||
~540ul of V4 probes: __ ng/ul => ug | |||
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===Qia Column Purification=== | ===Qia Column Purification=== |
Revision as of 20:38, 25 March 2015
CA12k_Nov2014_V4 Probe Production
- Production of Padlock Probes (V4) Matt:LabNotes/2014-11-19#Assembly_of_oligo_pool_for_ordering
- Do double 96 100ul PCR Reactions
Production PCR
- 10nM 1st round amplicons
- V4 master mix made in 2 5ml tubes
Components | Volume (1X) | Volume (100X) |
First round amplicon CA12k_Nov14_V7 (10nM) | 0.2 | 20 |
2X KAPA SYBG MM | 50 | 5000 |
AP1V4U (100uM) | 0.4 | 40 |
AP2V4 (100uM) | 0.4 | 40 |
H2O | 49 | 4900 |
Total | 100 | 10000 |
Program 98C 1min -> (98C 10sec -> 55C 30sec -> 72C 45sec) x 15 -> 72C 2min -> 15C hold File:032515 CA12kNov14 ProductionPCR V4.JPG
Zymo DNA Clean & Concentrator 100
- Purify 10ml of PCR product with 1 Zymo-spin V Column/Reservoir
- Add 50ml DNA Binding Buffer to 10ml product (Use 2 50ml tubes)
- Elute with 540ul H2O
- Measured concentration with Nanodrop:
~540ul of V4 probes: __ ng/ul => ug