id	author	title	date	pages	extension	mime	words	sentences	flesch	summary	cache	txt
cord-010329-rcx450b6	Khazak, Vladimir	Development of a Yeast Two-Hybrid Screen for Selection of Human Ras-Raf Protein Interaction Inhibitors	2005		.txt	text/plain	3549	184	51	The methods outlined below describe (1) the construction of expression plasmids bearing conditionally regulated yeast HXT9 and HXT11 hexose transporter genes, (2) integration of HXT9-and HXT11-containing DNA fragments in the PDR1 and PDR3 chromosomal loci of yeast two-hybrid strains, (3) analysis of the permeability of newly developed yeast strains with selected known smallmolecule inhibitors, and by screen of a large combinatorial library of compounds, and (4) selection of inhibitors of the interaction between human Ras and Raf-1 by screening the combinatorial library of compounds in the obtained hyperpermeable yeast two-hybrid strain. cerevisiae to small-molecular-weight compounds, the two yeast hexose transporters HXT9 and HXT11 were subcloned under control of the galactose-inducible GAL1 promoter and subsequently integrated by homologous recombination into the genetic loci for PDR1 and PDR3, thereby destroying the coding sequence of these genes. The plasmid pPDR3-HisInt was digested with AatII-SacI and the purified integrative cassette, hisG-URA3-PDR3-Int (see Fig. 2C ) was transformed into parental strains SKY48 and SKY191.	./cache/cord-010329-rcx450b6.txt	./txt/cord-010329-rcx450b6.txt