From 2008.igem.org
(Difference between revisions)
|
|
Line 40: |
Line 40: |
| </html> | | </html> |
| | | |
- | http://openwetware.org/images/thumb/6/61/UW_A%2BT.jpg/350px-UW_A%2BT.jpg http://openwetware.org/images/thumb/c/c7/UW_AT.jpg/350px-UW_AT.jpg
| + | |
| | | |
| | | |
Revision as of 09:21, 11 October 2008
|
|
|
|
Preparation of pMPMT5-AID+T7 and pMPMT5+AID-T7
Michał K.:
- Digest of PCR product for transcription fusion (T7 RNA-polymerase) with HindIII and SalI (2x Tango buffer).
- Digest of PCR product for translation fusion (AID+T7 RNA-polymerase) with NcoI and XbaI (1x Tango buffer).
- Clean-up of digested products from p.1 and p.2.
- Digest of pMPM-T5+AID with HindIII and SalI (2x Tango buffer), dephosphorylation (CIAP).
- Digest of pMPM-T5 with NcoI and XbaI (1x Tango buffer), dephosphorylation (CIAP).
- Gel-electrophresis and gel-out of proper bands (p.4 - 4850 bp, p.5 - 4250 bp).
- Electrophoresis to estimate the concentration of purified DNA.
- Ligation of purified products from p.1 and p.4.
- Electroporation of E. coli TOP10 with ligation product.
- Ligation of digested products from p.4 and p.5.
- Electroporation of E. coli TOP10 with ligation product.
- Transformants plating on LB + tetracycline.
Rifampicin test
Michał L., Ewa, Marcin:
- Plating colonies from the previous day on rifampicin.
|
|