Team:DTU Denmark/parts

C-terminal domain of Saccharomyces cerevisiae cyclin 2 (CLN2) gene. It has been shown that this region of the protein, which is rich of PEST motifs, leads to a destabilization of the protein. Hence this Tag can be used to increase the turn-over rate of a protein. In the article by Mateus and Avery: "Destabilized green Fluorescent protein for monitoring dynamic changes in yeast gene expression with flow cytometry" they show that addition of these 178 carboxyl-terminal amino acid residues changes the half-life of a GFP from 7h and down to 30 minutes.

Green Fluorescent Protein (GFP) codonoptimized for yeast.

This variant of biobrick BBa_K194001 has been destabilized by fusing biobrick BBa_K194000. In the article by Mateus and Avery: "Destabilized green Fluorescent protein for monitoring dynamic changes in yeast gene expression with flow cytometry" they show that addition of these 178 carboxyl-terminal amino acid residues of the Cln2 PEST signal, changes the half-life of a GFP from 7h and down to 30 minutes.

This biobrick contains a PacI/Nt.BbvCI and a AsiSI/Nb.BtsI USER cassette for insertion of PCR fragments using the USER cloning standard. PCR fragments containing uracil is treated with a uracil DNA glycosylase that removes the uracil exposing a predesigned 8bp overhang allowing for cloning without the need for ligase.