Hosuk:LabNotes/2014-3-13
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Mutation Detection with Mixed with PGP1F and NCIH1975 : Ampligase, 2nd RCA
- Continied from 03/06
Experiment
- 8 Ampligase reaction,
- 4 NTC(no Ampligase emzyme)
File:Mix PGP1F NCIH1975 24wellPlates AmpligaseMap.png
- Procedure
- Make Ampligase mix, add
- Incubate at 45C for 24hr. (03/13 ~ 03/14)
- Run RCA reaction for 24hr. (03/14 ~ 03/15)
- Anneal dcProbes, and Image
- Ampligase Mix
- 10x Buffer 20uL
- Ampligase (5U/uL) 8uL
- ppEGRF_T790M_WT 2uL
- ppEGRF_T790M_MU 2uL
- ppEGRF_L858R_WT 2uL
- ppEGRF_L858R_MU 2uL
- H2O 164 uL
- NTC Mix
- 10x Buffer 20uL
- Ampligase (5U/uL) 0uL
- ppEGRF_T790M_WT 2uL
- ppEGRF_T790M_MU 2uL
- ppEGRF_L858R_WT 2uL
- ppEGRF_L858R_MU 2uL
- H2O 172 uL
Result : 1st Rolonies
- Number of rolonies from most of wells are not bad.
- PGP1F has more 1st rolonies than cancer cell has.
- I think main reason is the size of PGP1F is much more than cancer cell's, which make a big different.
- That means the density per area of rolonies seems similar at each cell type.
File:Mix PGP1F NCIH1975 Counts.png
File:Mix PGP1F NCIH1975 Well04.png