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===Antibiotic=== | ===Antibiotic=== | ||
==Types of Screening== | ==Types of Screening== | ||
[[Image:Blue white test.jpg|thumb|right|Successful example of a blue/white screen test. Blue colonies are wild-type cells, while white colonies are successfully transformed cells]] | [[Image:Blue white test.jpg|thumb|right|Successful example of a blue/white screen test. Blue colonies are wild-type cells, while white colonies are successfully transformed cells]] | ||
===Green Fluorescent Protein=== | ===Blue/White Screening=== | ||
Blue/White Screening is commonly used in E. coli transformations. In this screening, cells are grown on agar plates in the presence of X-gal and IPTG to test for the presence of β-galactosidase enzyme. In the M15 strain of E. coli, part of the <i>lacZ</i> gene is deleted, removing the cell's ability to produce β-galactosidase. However, when transfected with a plasmid containing a <i>lacZα</i> domain, such as pUC19, the gene becomes operable and the cell produces β-galactosidase. It is possible to create a successful transformation in which β-galactosidase is not produced by inserting DNA into the <i>lacZα</i> domain. This is particularly useful to check for successful ligations. Successful ligations will not produce β-galactosidase, while unsuccessful ligations will. | |||
X-gal, while normally colorless (i.e. white), will readily hydrolyze in the presence of β-galactosidase into a compound with a sharp blue color. Therefore, colonies with successfully transformed cells with the desired DNA will grow white, while background colonies will grow blue. | |||
===Green Fluorescent Protein Screening=== | |||
[[Image:Green_Fluroescent_Mice.jpg|thumb|left|Mice transfected with GFP. One can easily distinguish the wild-type mouse (middle) from the two mice with GFP (left and right)<cite>Moen2011</cite>.]] | [[Image:Green_Fluroescent_Mice.jpg|thumb|left|Mice transfected with GFP. One can easily distinguish the wild-type mouse (middle) from the two mice with GFP (left and right)<cite>Moen2011</cite>.]] | ||
Green Fluorescent Protein, or GFP, was first isolated from the crystal jellyfish Aequorea victoria in the 1960s. In 1994, GFP was successfully cloned<cite>Chalfie1994</cite>, allowing researchers to use the protein as a screenable marker for the first time. Virtually harmless in live cells, GFP has the unique pheotype of glowing bright green under ultraviolet light. GFP functions entirely of its own accord, and requires no exogenous material besides ionizing radiation in order to fluoresce. This allows GFP to be used as a marker accompanying transfected DNA, and has been used extensively in academia. | Green Fluorescent Protein, or GFP, was first isolated from the crystal jellyfish Aequorea victoria in the 1960s. In 1994, GFP was successfully cloned<cite>Chalfie1994</cite>, allowing researchers to use the protein as a screenable marker for the first time. Virtually harmless in live cells, GFP has the unique pheotype of glowing bright green under ultraviolet light. GFP functions entirely of its own accord, and requires no exogenous material besides ionizing radiation in order to fluoresce. This allows GFP to be used as a marker accompanying transfected DNA, and has been used extensively in academia. | ||
