Glasgow/Wetlab

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PROTOCOLSREFERENCES

Contents

Week 1

03/07

  1. Maija and Christine prepared LB broth and LB agar with Protocol 1. Made 4 batches of plates, 2 with kanamycin and 2 with carbenicillin.

04/07

  1. All wetlab researched BioBricks.
  2. Reporter constructs and mini-Tn5 stocks looked out.
  3. Streaked the following:
    1. P. putida PAW 340 pJAK14 (Carb. plate)
    2. Tn5 lux AB (Carb. plate)
    3. Mini-Tn5 lux AB (Carb. plate)
    4. P. Fluorescens NCIMB 9815 (Carb. plate)
    5. P. putida KT2440 (LB)
    6. JM109 pBluescript 5k+ (Carb. plate)
    7. Mini-Tn5 Tc (Carb. Plate)
    8. pQF52 (Carb. plate)
    9. P. Fluorescens 9815 (LB)
    10. E. coli pJAK14 (Km plate)
    11. Il DntR in pOF52 (Carb. plate)
    12. pUCINR in Ω strain C (Carb. plate)
    13. pGLTUR (Carb. plate)
    14. Mini-Tn5 Kan (Carb. plate)
    15. Mini-Tn5 Sm/Sp (Carb. plate)
    16. Mini-Tn5 1cc2 (Carb. plate)
    17. E. coli Sa1 (LB)
    18. DmpR #24 (Carb. plate)
    19. Mini-Tn5 lac 32 in E. coli 517 (Carb. plate)
    20. Mini-Tn5 Tc (Carb. Plate)
    21. Mini-Tn5 Cm (Carb. plate)
    22. DmpR WT (Carb. plate)
    23. E.coli sm 10 pESD15 Tn5 GFP (Carb. plate)
    24. pUJ8 (Carb. Plate)

05/07

  1. BioBricks – Maija and Scott transformed using Protocol 2.
    • BBa_p1010 (DB3.1) (death gene plasmid)

Plate 4:
7A – p5B1A10
11E – p5B1A10
11C – p5B3K3

    • BBa_IS2001 (Top10) (high copy number plasmid)

3 Plate 4: 5I – p5B4A5 4 5D – p5B4K5 5 6B – p5B3K5 6 6D – p5B4K5 7 6E – p5B4A5 8 (Also 5K, 5M, 4J, 4L, 4N, 4P) 9 BBa_J23119 (Top 10) (strong constitutive promoter) 10 Plate 3: 19A – pB1A2 (V1013) 11 BBa_R0062 (Top 10) (HSL and luxR inducible) 12 Plate 1: 9G – pSB1A2 (V1004) 13 BBa_J04500 (Top 10) (IPTG inducible prom + RBS) 14 Plate 1: 16P – p5B1AK3 (V1009) 15 BBa_E0040 (GFP mutant no promoter (3b)) 16 Plate 1: 5H – p5B1A2 (V1001) 17 Maia and Christine researched restriction enzymes for the reporter constructs and Mini-Tn5s streaked on 4/7/07 and digested some. Had planned to make biobricks using Mini-Tn5s etc but discovered that a previous attempt to use transposable elements to make biobricks had been unsuccessful due to scarring at the restriction site (http://partsregistry.org/Part:BBa_J61206).