Noi/NOTES/2014-8-21: Difference between revisions
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>Noi (Created page with "* '''Link to calendar''' * 2014-08-13: Received MOMOD V4 Oligo pool from CustomAraay === Oligo info. === * Name: 90K_ologos_04Aug2014 (8-20-14) * Con...") |
>Noi |
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== Expansion PCR (Quick test) == | == Expansion PCR (Quick test) == | ||
* I did a quick test of expansion PCR on the three subset. | * I did a quick test of expansion PCR on the three subset. | ||
* I diluted 1st amplicon of each primer set ( | * I diluted 1st amplicon of previous probe set amplified by each primer set (by V4, V6, or eMIP_CA primers) into 2nM and will be as positive control for amplification. | ||
** V4 set: I used MONOD GP1V4 as template. Mix 10ul of 20nM 1st round amplicon MONOD GP1V4 with 90ul H2O | ** V4 set: I used MONOD GP1V4 as template. Mix 10ul of 20nM 1st round amplicon MONOD GP1V4 with 90ul H2O | ||
** V6 set: I used MONOD V3 as template. Prepare 2nM template in the same way as V4 set. | ** V6 set: I used MONOD V3 as template. Prepare 2nM template in the same way as V4 set. |
Revision as of 01:40, 22 August 2014
- Link to calendar
- 2014-08-13: Received MOMOD V4 Oligo pool from CustomAraay
Oligo info.
- Name: 90K_ologos_04Aug2014 (8-20-14)
- Conc. 153.49ng/ul,
- Volume 80ul in TE buffer. Total amount 12.28ug
- Info from Dr. Zhang's note
Description # oligos Length Amp. Primers Adaptor Removal Set A: BSPP LMS cluster probes 73,574 148nt V6 USER/DpnII Set B: Selector LMS cluster 12,994 148nt V4 USER/DpnII Set C: Selector chr21 unique 2,539 148nt eMIP_CA_F/R Nt.AlwI/Nb.BrsDI Total: 89,107
- I will be working on 73K probe set
- Length 148 mer (MW = 148 * 330Da/bp = 48,840 Da or g/mole
- Conc. 20.53ng/ul = 3.1427uM or 3142.7nM (153.49ng/ul/ 48,840g/mole)
Expansion PCR (Quick test)
- I did a quick test of expansion PCR on the three subset.
- I diluted 1st amplicon of previous probe set amplified by each primer set (by V4, V6, or eMIP_CA primers) into 2nM and will be as positive control for amplification.
- V4 set: I used MONOD GP1V4 as template. Mix 10ul of 20nM 1st round amplicon MONOD GP1V4 with 90ul H2O
- V6 set: I used MONOD V3 as template. Prepare 2nM template in the same way as V4 set.
- eMIP_CA: I used oligo from LC sciences used in Nature methods 2012. Mix 20ul of 10nM 1st round amplicon 4K LC Sciences with 80ul H2O
95C 30sec -> (95C 30sec -> 54C 45sec-> 72C 45sec) X 18-> 72C 3min -> 15C hold