Team:Tianjin/Note/Consortium

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Team Tianjin-Attribution

Week2(7/31/2016-8/6/2016)

  • optimization of culture conditions


      Aug.4th

            Observed the growth of 1-day pre-cultured bacteria in LB at 30℃:
    All above were kept culturing for one more day and were checked on the next day. The next step was to explore the optimal condition for each bacterium and to co-culture each teo of them to see whether the consortium could work well.

    The most probable pairs:
    1. P.putida KT2440 + R.jostii RHA1;
    2. B.subtilis 168 + R.jostii RHA1;
    3. B.subtills 168 + P.putida KT2440;
    4. R.jostii RHA1 + Y.lipolytia;
    5. P.putida KT2440 + Y.lipolytia;
    6. E.coli(RFP) + Y.lipolytia.

      Aug.5th

    Cultured different bacteria in M9 medium with sodium terephthalate at 30℃ & 200 rpm, checked the bacterial concentration at OD600 and detected the concentration of TPA by UV at OD242: The growth of each bacterium was not as expected, so we considered culturing bacteria in improved W medium.

  • Construction of PBBR


      Aug.5th

            Extraction of plasmid pBBR1MCS-2

      Aug.6th

                Cut pBBR1MCS-2 with restricted enzymes Xba1 and Sac1 and checked by agarose gel electrophoresis:
            Failed. After gel recovery, the concentration of digested pBBR1MCS-2 was 2.4 ng/μl, which was too low to be used in the next step.
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    Week5(8/21/2016-8/27/2016)

  • optimization of culture conditions


      Aug.22th

            1.Prepared 200ml W0 medium and add 0.3g NaOAc, then regarded it W7 medium.
            2.Extractd 8ml W7 medium to 16 test tubes, respectively.
            3.Added becteria solution as following table (use two tubes each group)
          4.cultured them at 30℃ and check growing situation and the situation of TPA degradation.
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    Week6(8/28/2016-9/3/2016)

  • optimization of culture conditions


      Sep.12th

    Optimize the growing environment of Bacillus stubtilis by change medium components. Firstly, we prepared 400ml W medium, then, we devided the medium into four pieces averagely and number them No.1, No.2, No.3, No.4. Next, we added 0.1g NaCl, 0.1g NH4Cl, 0.3g sucrose to No.2, No.3, No.4, respectively. After that, we extracted 5 ml No.1, No.2, No.3, No.4 and added them to four test tubes, then, we added 10 μL bacteria solution of Bacillus stubtilis 168 to four test tubes, respectively. Eventually, we cultured them at 37℃ and check growing situation.
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    Week8(9/11/2016-9/17/2016)

  • optimization of culture conditions


      Sep.12th

    Optimize the growing environment of Bacillus stubtilis by change medium components.Firstly, we prepared 400ml W medium, then, we devided the medium into four pieces averagely and number them No.1, No.2, No.3, No.4. Next, we added 0.1g NaCl, 0.1g NH4Cl, 0.3g sucrose to No.2, No.3, No.4, respectively. After that, we extracted 5 ml No.1, No.2, No.3, No.4 and added them to four test tubes, then, we added 10 μL bacteria solution of Bacillus stubtilis 168 to four test tubes, respectively. Eventually, we cultured them at 37℃ and check growing situation.

    •   Sep.16th

            1.Prepared 200ml W0 medium and add 0.2g NH4Cl, then regarded it W8 medium.
            2.Extractd 10ml W8 medium to 16 test tubes, respectively.
            3.Added becteria solution as following table (use two tubes each group)
            4.cultured them at 30℃ and check growing situation and the situation of TPA degradation.
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    Week9(9/18/2016-9/24/2016)

  • optimization of culture conditions


      Sep.19th

            1. Prepared 1L W medium and add 2.5g TPA and 1.1875g NaOH.
            2. devided the medium into five pieces averagely and add chemicals as following table
            3. Extractd 5ml W9, W10, W11, W12, W13 medium to 40 test tubes, respectively.
            4. Added becteria solution as following table (use two tubes each group)
            5. cultured them at 37℃ and check growing situation.


      Sep.21th

            Repeat experiments of W9, W10 the day before yesterday.


      Sep.22th

            1. Use five mediums of Sep.19th and W0, W8 medium of Sep.16th.
            2. Extractd 5ml W0, W8, W9, W10, W11, W12, W13 medium to 56 test tubes, respectively.
            3. Added becteria solution as following table (use two tubes each group)
            4. cultured them at 37℃ and check growing situation.


      Sep.23th

            1. Prepared 400L W medium and add 1.2g KNO3 and 1.2g glucose.
            2. devided the medium into four pieces averagely and add chemicals as following table
            3. Extractd 5ml W0, W14, W15, W16 medium to 34 test tubes, respectively.
            4. Added becteria solution as following table (use two tubes each group)
            5. cultured them at 30℃ and check growing situation.
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