TY  - JOUR
A1  - Miyamoto-Sato, Etsuko
A1  - Ishizaka, Masamichi
A1  - Horisawa, Kenichi
A1  - Tateyama, Seiji
A1  - Takashima, Hideaki
A1  - Fuse, Shinichiro
A1  - Sue, Kaori
A1  - Hirai, Naoya
A1  - Masuoka, Kazuyo
A1  - Yanagawa, Hiroshi
T1  - Cell-free cotranslation and selection using in vitro virus for high-throughput analysis of protein–protein interactions and complexes
Y1  - 2005/05/01 
JF  - Genome Research 
JO  - Genome Research 
SP  - 710 
EP  - 717 
DO  - 10.1101/gr.3510505 
VL  - 15 
IS  - 5 
UR  - http://genome.cshlp.org/content/15/5/710.abstract 
N2  - We have developed a simple and totally in vitro selection procedure based on cell-free cotranslation using a highly stable and efficient in vitro virus (IVV). Cell-free cotranslation of tagged bait and prey proteins is advantageous for the formation of protein complexes and allows high-throughput analysis of protein–protein interactions (PPI) as a result of providing in vitro instead of in vivo preparation of bait proteins. The use of plural selection rounds and a two-step purification of the IVV selection, followed by in vitro post-selection, is advantageous for decreasing false positives. In a single experiment using bait Fos, more than 10 interactors, including not only direct, but also indirect interactions, were enriched. Further, previously unidentified proteins containing novel leucine zipper (L-ZIP) motifs with minimal binding sites identified by sequence alignment as functional elements were detected as a result of using a randomly primed cDNA library. Thus, we consider that this simple IVV selection system based on cell-free cotranslation could be applicable to high-throughput and comprehensive analysis of PPI and complexes in large-scale settings involving parallel bait proteins. 
ER  -