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|Grow'n'Glow GFP Yeast One-Hybrid System. The Grow'n'Glow GFP One-Hybrid kit isolates genes for proteins that bind a specific DNA element of interest. In addition to finding novel DNA-binding proteins, the one-hybrid system can be used to investigate the bases and amino acids involved in specific DNA-protein interactions. Proteins can be found that bind to any short DNA element of interest. The Grow'n'Glow system offers maximal sensitivity because detection of the DNA-protein interaction occurs in vivo, where proteins are more likely to be in their native conformation. Use of the Green Fluorescent Protein (GFP) reporter eliminates the time-consuming beta-galactosidase lift assays. The particular GFP variant expressed by the GFP reporter plasmid, GFPuv, has the same excitation and emission maxima as wild-type GFP, but is 18 times brighter than the wild-type variant. Positive colonies with DNA-binding proteins glow brightly green and can be easily detected by placing the plate with the yeast colonies on a standard 300 nm UV transilluminator or under a UV lamp in a darkroom. Expression of fusion proteins by the prey plasmid is controlled by the GAL1 upstream-activation sequence. In yeast with an intact galactose regulatory system, the GAL1 activation sequence is induced by galactose and repressed by glucose. This regulation inhibits expression of library/activation domain fusions until the actual screening, which prevents any potentially toxic library proteins affecting growth of the yeast. This sort of toxic proteins would be missed during screening in a non-inducible system. Features: • Isolates genes for novel DNA-binding proteins • Maps residues and regions responsible for DNA-binding • Positive colonies fluoresce brightly green • One-step selection of potential positives under UV light • No time-consuming beta-galactosidase lift assays as essential in most conventional assays • Finds potentially toxic proteins by screening with galactose-inducible expression libraries
|mixed; -80° C and 4° C
|Two- and One-Hybrid Systems