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Hello
Yes, welcome to our blog! remember to wipe your legshands with ethanol before coming in. Don't leave any bacteria behind. We are currently in the Molecular Biotechnology & learning about microorganisms.

=D

Group Members

  • Sairam (leader)
  • Fang Cheng
  • Yan Ping aka Malau
  • YiJu
  • Edyna
  • Wileen
  • Wan Rong
  • Ida
  • Zhiyuan aka Pig
  • Kelvin
  • E-young

  • Links

    edyna @ blogspot
    wanrong @ blogspot
    yitian @ blogspot
    xiao bin @ blogspot

    Previous Post

    How GFP gene is incorporate in E.Coli?
    Video on handling the setup
    Day 1
    Day 2
    Day 3
    Day 4

    History

    November 2007

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    Tuesday, November 6, 2007
    Theory behind GFP ( Pre-Practical)

    Green Fluorescent Protein (GFP) has existed for more than one hundred and sixty million years in one species of jellyfish, Aequorea victoria. It is a protein produced by Aequorea victoria which fluoresces green when exposed to blue light, and it is found in the photoorgans of Aequorea victoria. Upon binding aequorin, a protein in Aequorea victoria with calcium, blue light is released and then totally absorbed by the GFP giving off green light.


    Fig 1: Aequorea victoria
    Fig 2: Aequorea victoria photoorgans

    (Pictures taken from
    http://www.conncoll.edu/ccacad/zimmer/GFP-ww/GFP-1.htm)

    GFP comprises 238 amino acids (26,9 kDa) and takes on a unique can-like shape consisting of an 11-strand β-barrel with a single alpha helical strand containing the chromophore running through the center. The inward facing sidechains of the barrel induce specific cyclization reactions that lead to chromophore formation, while the tightly packed barrel shell protects the chromophore from quenching by the surrounding microenvironment. Chromophore formation occurs in a series of discrete steps with distinct excitation and emission properties (maturation).

    GFP does not require exogenous moiety for fluorescence. It is a naturally fluorescent protein in which the chromophore (fluorophore) is derived from post-translational cyclization of a serine-tyrosine-glycine tripeptide of GFP, followed by dehydrogenation of the tyrosine. This makes it a tremendously useful marker in in vivo studies. In fact, since its original discovery in the jellyfish Aequorea victoria, it has proven valuable in a plethora of biochemical, cellular, and developmental investigations. It is now found in almost all the laboratories in all over the world where it is used in every conceivable plant and animal. Organisms such as E. coli, flatworms, algae, and pigs have all been made to fluoresce with GFP.

    GFP are usually much less harmful when illuminated in living cells compared to most of the small fluorescent molecules such as FITC (fluorescein isothiocyanate). This triggered the development of highly automated live cell fluorescence microscopy system for observation of cells over time expressing one or more proteins tagged with fluorescent proteins. Understanding of many biological processes such as protein folding, protein transport, and RNA dynamics, which in the past had been studied using fixed (i.e. dead) material has then been redefined with the analysis of such time lapse movies.

    GFP is like the microscope of the twenty-first century where people use it to see when proteins are made and where they can go. This can be done by joining the GFP gene to the gene of the protein of interest so that when the protein is made it will have GFP hanging off it. Light can be shine at the cell and wait for the distinctive green fluorescence associated with GFP to appear. One may thus have an in vivo fluorescent protein which may be followed in a living system. There have been several recent developments for the use of GFP and several different colour variants.

    Expression of the protein in small sets of specific cells is another powerful use of GFP, allowing researchers to optically detect specific types of cells in vitro (in a dish), or even in vivo (in the living organism). The GFP gene can be introduced into organisms and maintained in their genome through local injection with a viral vector and breeding. Many bacteria, yeast and other fungal cells, plant, fly, and mammalian cells have been created using GFP as a marker.



    Fig 3: Visually arresting photograph that have been taken of fluorescently labelled proteins.

    (Picture taken from http://www.conncoll.edu/ccacad/zimmer/GFP-ww/GFP-1.htm)



    7:36 AM


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