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Team:Bielefeld-CeBiTec/Notebook/Journal/Isobutanol/Oct
From 2014.igem.org
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October |
- pSB1C3_T7_adhA
- This week we tried to reclone pSB1A2_T7_adhA into the pSB1C3 backbone.
- Because the recloning didn't worked well till now, we decided to restrict the used backbone pSB1C3_RFP with EcoRI, XbaI, SpeI and PstI to minimize the chance of religation.
- pSB1C3_T7_adhA we cut with EcoRI and PstI
- Clean up and Ligation as explained in the BioBrick Assembly
- Transformation with electrocompotetent cells
- Colony PCR (rev_pB1C3_adhA, fw_adhA_pSB1C3)
- Annealing temperature: 65 °C
- Bands as expected (~1200 bp)
- Liquid cultures for a restriction digest were prepared.
- Plasmid isolation of pSB1C3_T7_adhA
- Restriction digestion with EcoRI and PstI
- Bands as expected (backbone: ~2,2 kb and insert: ~1,2 kb)
