Team:UGA-Georgia/Notebook

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<a href="https://2014.igem.org/Team:UGA-Georgia"><p style="font-family: Basic L"><font size="2">HOME</font></p> </a> </td>
<a href="https://2014.igem.org/Team:UGA-Georgia"><p style="font-family: Basic L"><font size="2">HOME</font></p> </a> </td>
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         onmouseout="mclosetime()" style="color:#000000"><p style="font-family: Basic L"><font size="2"><b>HUMAN PRACTICES</b></font></p></a>
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Revision as of 02:35, 17 October 2014



HOME

PROJECT

WET LAB

HUMAN PRACTICES

TEAM

Notebook

February


Week 1
  • Introduction to anaerobic facilities for new members

    • Week 3
  • Making of NaS solution.
  • Making of formate media.

    • Week 4
  • Creation of transformation buffer (TB).
  • First geraniol extraction efficiency test with balch tubes.


    • March


    Week 1
  • Creation of geraniol standards for gas chromatography (GC) analysis/calibration through serial dilutions.

    • Week 3
  • Creation of more formate media.
  • Creation of general salts solution.
  • Creation of glycylglycine buffer.


    • April


    Week 1
  • Learning about ribosome binding site library (RBS) and our metabolic Optflux model.

    • Week 2
  • Made agar plates
  • Creation of fresh wild type (WT) stocks.

    • Week 3
  • Created diluted puromycin


    • May


    Week 4
  • Inoculation and revival of frozen stocks.


    • June


    Week 1
  • Determined the optical density (OD) of the previously revived cultures.

    • Week 2
  • Restriction digestion, purification, and ligation of pMEV4 and PCR products (C1 and C2).
  • Transformation by heat shock, plated the cells, made an agarose gel and ran the products on it.
  • Picked colonies from the previous day of plating.
  • Plasmid extraction, digestion, gel electrophoresis, and verification.

    • Week 3
  • Revival of pAW50-mCherry frozen stock.
  • Revival of S0001 (WT) frozen stock.

    • Week 4
  • Prepared the revived cultures for fluorescence microscopy.
  • Fluorescence microscopy training.
  • Anaerobic transformation of RBS libraries 1-12 into M. maripaludis.
  • Puromycin enrichment of transformants.
  • Made glycerol stocks of original transformants.
  • Fluorescent microscopy of transformants/pAW50-mCherry/S0001.


    • July


    Week 1
  • Took ODs of the native RBS and library 1 and 2 (12; L-1; L-2 respectively).
  • Plated the 12, L-1, and L-2 cultures.
  • Dispensed media to test tubes.
  • Picked colonies form the plates.

    • Week 2
  • ODs from last weeks picked colonies were taken.
  • Made frozen stocks of RBS colonies,
  • Sub-cultured parent strains.
  • Researched primary literature on mCherry maturation.
  • Began developing a protocol for oxygen maturation of mCherry.

    • Week 3
  • Made frozen stocks of RBS colonies and sub-cultured parent strains.
  • Re-subcultured and revive pAW50-mcherry.
  • Subcultured the parent strains of WT and native RBS (12) into triplicates. The triplicates were wrapped in foil to prevent any photobleaching.
  • The pAW50-mcherry came up and then all samples were taken to a plate reader to perform fluorescence tests.

    • Week 5
  • Filled out and submitted safety form.
  • The geraniol extraction efficiency experiment was performed.
    • Left the organic phase to dry over night.

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