Team:Munich/thisisatest.html
Transforming E.Coli NEB Turbo to amplify plasmids for pRED/ET Engineering
2018/05/07| Participants: | Dominic Schwarz |
| Protocol: | Electro-transformation |
| Notes: | pkD3 contains resistance cassettes with FRT-sites for pRED engineering. Due to the temperature sensitive promotor pNPTS138-R6KT which is used for bone-knockin via RecA recombineering, the cells were incubated at 30°C overnight. |
| Results: | No colonies. We suspected electroporation to be a problem and switched to chemical transformation in NEB Turbo next. |
Redo: Transforming E.Coli NEB Turbo to amplify plasmids for pRED/ET Engineering
2018/05/18| Participants: | Dominic Schwarz |
| Protocol: | Chemical Transformation |
| Notes: | Inoculate pRED at 30°C because of temperature sensitive promoter |
| Results: | No colonies. We suspected the E. Coli NEB Turbo to be a problem and switched to E. Coli Dh5α. |
Redo: Transforming E.Coli Dh5α to amplify plasmids for pRED/ET Engineering
2018/05/24| Participants: | Dominic Schwarz |
| Protocol: | Electro-transformation |
| Notes: | inoculate pRED at 30°C because of temperature sensitive promoter, pKD3 contains resistance cassette flanked by FRT sites |
| Results: | We got cells containing the plasmid from PD Dr. Jürgen Lassak of the LMU. |
Transforming E.Coli Dh5α to amplify plasmids for pRED/ET Engineering
2018/05/25| Participants: | Dominic Schwarz |
| Protocol: | Chemical transformation |
| Notes: | pKD3 contains resistance cassette flanked by FRT sites |
| Results: | No colonies. Because pRED/ET could not be transformed, we got readily transformed cells from PD Dr. Jürgen Lassak from the LMU |
DNA preparation for pRED/ET Engineering
2018/05/26| Participants: | Dominic Schwarz |
| Protocol: | Miniprep |
| Notes: | The cells from PD Dr. Jürgen Lassak were used for DNA preparation. |
| Results: | Concentration of the plasmids obtains from MIniprep: pRED/ET: 37,5 ng/µl pNPTS138-R6KT: 60ng/µl |