Team:Bielefeld-CeBiTec/Notebook/Journal/CO2-fixation/Aug

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                 <h6><a  style="font-size:24px" href="#">Week 1 &nbsp;&nbsp;&nbsp; 08/04 - 08/10</a></h6>
                 <h6><a  style="font-size:24px" href="#">Week 1 &nbsp;&nbsp;&nbsp; 08/04 - 08/10</a></h6>
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                 <ul> <!-- gesamte Liste -->
                 <ul> <!-- gesamte Liste -->
                   <li><b>Promotors T7, p<sub>Tac</sub> and p<sub>Tet</sub></b></li>
                   <li><b>Promotors T7, p<sub>Tac</sub> and p<sub>Tet</sub></b></li>

Revision as of 14:35, 26 August 2014


August

Week 1     08/04 - 08/10
Week 1     08/04 - 08/10
  • Promotors T7, pTac and pTet
    • We tried to assmeble some inserts with three different promotors to test which one is the best choice.
      • BioBrick Assembly (Suffix)
        • Backbones (digested with SpeI, PstI)
          • T7
          • pTac
          • pTet
        • Inserts (digested with XbaI, PstI)
          • prkA
          • Hneap
          • SELAN
          • sRNA:pfkA
        • Only the assembly with the pTac promotor was successful.
      • Colony PCR and plasmid isolation of pTac_prkA, pTac_Hneap, pTac_SELAN and pTac_sRNA_pfkA
      • Colony PCR of T7 promotor
        • Showed in all cases an band 400 bp over the expected value. We tried to extract and tranform the promotor from another distribution plate (2013)
      • Colony PCR of T7 promotor from the 2013 distribution plate
        • Bands as expected (~300 bp)
  • csoS1-4
    • We tried to [...]
      • Colony PCR (VF-Primer, VR-Primer)
      • Gibson Assembly
      • Plasmid isolation
      • Restriction digestion with SpeI and XbaI
        • Bands as expected ()
      • BioBrick Assembly (Suffix)
        • Backbone (digested with SpeI, PstI)
          • pSB1C3_can
        • Insert (digested with XbaI, PstI)
          • csoS1-4
      • Colony PCR of can_csoS1-4
        • Bands as expected ()
  • glpX
    • We tried to [...]
      • Gibson Assembly after DpnI digestion
      • Colony PCR
  • prkA
    • We tried to [...]
      • PCR amplification and purification for insertion in pHnCBcsoS1D
        • Bands as expected ()
      • Gibson Assembly with pHnCBcsoS1D
  • RuBisCO
    • We tried to...
      • BioBrick Assembly (Suffix)
        • Backbone (digested with SpeI, PstI)
          • pTac_prkA
        • Inserts (digested with XbaI, PstI)
          • Hneap
          • SELAN
        • We assembled pTac_prkA with Hneap respectively SELAN
      • Colony PCR
        • pTac_prkA_Hneap
          • Bands es expected ()
        • pTac_prkA_SELAN
          • Bands as expected ()
      • Plasmid isolation of pTac_prkA_Hneap and pTac_prkA_SELAN
  • Sequencing
    • csoS1D
    • Successful. We got the right sequence with 100%. The first part of the carboxysome is complete.
    • can
    • Five mutations. Another sequencing will follow.
    • glpX
    • Not successful. We got the CFP sequence instead of the glpX sequence, so we will try another backbone (pSB1C3_RFP).

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