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Team:HokkaidoU Japan/Notebook/aggregation Week 9
From 2012.igem.org
August 27th
Colony PCR
Colony PCR to confirm that whether the pT7-RBS-Ag43-dT on pSB1C3 was successfully ligated or not.
| DNA solution | 4 ul |
| Kapa-Taq(Taq polymerase) | 5 ul |
| Forward Primer(ag43-f4 primer) | 0.5 ul |
| Reverse Primer(200bp down primer) | 0.5 ul |
| Total | 10 ul |
| Number | Degree | Second |
| 1 | 95 | 120 |
| 2 | 95 | 30 |
| 3 | 53.0 | 30 |
| 4 | 72 | 60 |
| 5 | 72 | 60 |
| 6 | 4 | HOLD |
Cycle:2~4 x 35
We used N1 (DW only) and N2 (Ag43-dT on pSB1AK3) as controls. Desired product is about 695bp.
[[image:|thumb|Colony PCR result]]
Incubation for mini-prep
Incubation of pT7-RBS-Ag43-dT on pSB1C3 in LBC liquid medium.
- Prepared 2 ml LBC into culture tubes.
- Re-suspended 2 colonies (No.1 and No.2 respectively).
- Incubated at 37C for 15 hrs.
