Rapid selection against truncation mutants in yeast reverse two-hybrid screens

Citation
H. Puthalakath et al., Rapid selection against truncation mutants in yeast reverse two-hybrid screens, BIOTECHNIQU, 30(5), 2001, pp. 984
Citations number
11
Categorie Soggetti
Biochemistry & Biophysics
Journal title
BIOTECHNIQUES
ISSN journal
07366205 → ACNP
Volume
30
Issue
5
Year of publication
2001
Database
ISI
SICI code
0736-6205(200105)30:5<984:RSATMI>2.0.ZU;2-D
Abstract
The yeast reverse two-hybrid system is a powerful technique for isolating m utations in a protein that abolish its interaction with a known partner. Se lection is based on abrogation of growth suppression imposed when wild-type interactions confer 5-fluoroorotic acid (5-FOA) sensitivity to yeast cells . A laborious component of this system is to eliminate those mutations that cause protein truncation. By fusing the green fluorescent protein (GFP) to the C-terminus of a protein of interest, dynein light chain (LC8), we were able to rapidly isolate mutations that did not result in protein truncatio n.