Team:HokkaidoU Japan/Notebook/aggregation Week 10
From 2012.igem.org
September 3rd
Colony PCR of eCFP-RBS-pSB1A2
DNA solution | 4 ul |
Kapa-Taq(Taq polymerase) | 5 ul |
Forward Primer(EX-F primer) | 0.5 ul |
Reverse Primer(PS-R down primer) | 0.5 ul |
Total | 10 ul |
Number | Degree | Second |
1 | 95 | 120 |
2 | 95 | 30 |
3 | 68.9 | 30 |
4 | 72 | 60 |
5 | 72 | 60 |
6 | 4 | HOLD |
Cycle:2~4 x 35
We used N1 (DW only) and N2 (ptetR-RBS-eCFP-dT-pSB1A2) as controls. Desired product is about 776bp.
We confirmed that about 70% of ligated DNA formed our desired construct. We selected No.2 and 5 colony for incubation and store No.7 and 8 colony mixture at 4C.
Incubation of eCFP-RBS-pSB1A2 for mini-prep
- Prepared 2 ml LBA into culture tubes.
- Re-suspended 2 colony mixture (No.2 and No.5 respectively).
- Incubated at 37C for hrs.