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Revision as of 10:17, 17 July 2015


iGEM Kent 2015

Notebook

  • June,2015
    • Sun Mon Tue Wed Thu Fri Sat
                  1
      2 3 4 5 6 7 8
      9 10 11 12 13 14 15
      16 17 18 19 20 21 22
      23 24 25 26 27 28 29
      30            
  • July,2015
    • Sun Mon Tue Wed Thu Fri Sat
        1 2 3 4 5 6
      7 8 9 10 11 12 13
      14 15 16 17 18 19 20
      21 22 23 24 25 26 27
      28 29 30 31      
  • August,2015
    • Sun Mon Tue Wed Thu Fri Sat
              1 2 3
      4 5 6 7 8 9 10
      11 12 13 14 15 16 17
      18 19 20 21 22 23 24
      25 26 27 28 29 30 31



















Day 1 Wet lab 22/06/15

  • Made LB plates
  • Made LB broth
  • TIPS autoclaved

    • Day 2 Wet lab 23/06/15

  • Set up Top10 overnight, VS45 overnight, US348 overnight
  • Meeting with advisors to discuss progress

    • Day 3 Wet lab 24/06/15

  • Filter sterilise the buffer
  • 250ml no antibiotic broth, put top10 cells in
  • Incubate until optical density is 0.6
  • Miniprep PSB1C3 plasmid

    • Day 4 Wet lab 25/06/15

  • Transformation
  • Prepare SBC media
  • Digest of PSB1c3 from MS348

    • Day 5 Wet lab 26/06/15

  • Calculated competent cell efficiency
  • Agarose gel formation
  • Gel electrophoresis

    • Day 6 Wet lab 29/06/15

    Transforming linear PSB1C3

    Day 7 Wet lab 30/06/15

  • Miniprep of PSB1A3 plasmid
  • Gel electrophoresis of a large quantity of PSB1C3
  • Meeting with advisors to discuss progress

    • Day 8 Wet lab 01/07/15

  • Gel electrophoresis of the purified DNA extracted
  • Transformation of pSB1A3 circular plasmid to VS45 cells (competent)
  • Digest of all pSB1C3 plasmids

    • Day 9 Wet lab 02/07/15

  • Competent cells transformation efficiency
  • Mini-prep of 1MS340 to get pSB1C3 plasmid
  • Gel extraction of big digest
  • Quantification of the digest. Added sample buffer to each digest

    • Day 10 Wet lab 03/07/15

  • Transformation efficiency

    • Day 11 Wet lab 06/07/15

  • TOP10 cells containing pSB1A3 with limonene synthase were cultured overnight on AMP plates
  • Colonies of VS45 containing pSB1A3 were patched onto chloramphenicol plates
  • Overnight digest of miniprepped pSBIC3 using ECORI and PSTl

    • Day 12 Wet lab 07/07/15

  • Agarose gel of overnight digest - no bands were visible
  • Set up overnight cultures of pSBIC3 and pSBIA3 to be miniprepped the next day
  • Set up an overnight digest of pSBIC3

    • Day 13 Wet lab 08/07/15

  • Agarose gel of overnight digest - no bands were visible
  • Miniprep of pSBIA3 and pSBIC3
  • Overnight digest of pSBIA3 and pSBIC3

    • Day 14 Wet lab 09/07/15

  • Agarose gel of digested pSBIA3 and pSBIC3 - no bands were visible
  • Overnight cultures of MS349 containing pSBIA3 LIMS and MS340 containing pSBIC3 were set up, ready to be miniprepped

    • Day 15 Wet lab 10/07/15

  • Miniprep of plasmids pSBIA3 and pSBIC3

    • Day 16 Wet lab 13/07/15

  • Overnight cultures of MS349 containing pSBIA3 LIMS and MS340 containing pSBIC3 were set up, ready to be miniprepped
  • Overnight digest of miniprepped plasmids pSBIA3 and pSBIC3 using the enzymes ECORI and PSTI

    • Day 17 Wet lab 14/07/15

  • Agarose gel of digested plasmids pSBIA3 and pSBIC3 - it finally worked!!!
  • Miniprep of overnight cultures
  • Gel extraction of both plasmids
  • Analytical gel using Hyperladder I to quantify the plasmid
  • Overnight cultures of Top10 cells containing PVS72

    Day 18 Wet lab 15/07/15

  • Miniprepped PVS72
  • Transformation of VS45 with PVS72
  • Made LB plates with Chloramphenicol and Ampicillin
  • Plated the transformed VS45 cells on the Chloramphenicol/Ampicillin plates
  • Agarose gel of leftover digested pSBIA3 and pSBIC3 from 14/07/15
  • Gel extraction of pSBIA3 and pSBIC3

    • Day 19 Wet lab 16/07/15

  • Counted the overnight colonies of VS45 with PVS72
  • Calculated the transformation efficiency