BAC TransgeneOmics:: a high-throughput method for exploration of protein function in mammals

被引：186

作者：

Poser, Ina ^{[2
]}

Sarov, Mihail ^{[1
,2
]}

Hutchins, James R. A. ^{[3
]}

Heriche, Jean-Karim ^{[4
]}

Toyoda, Yusuke ^{[2
]}

Pozniakovsky, Andrei ^{[2
]}

Weigl, Daniela ^{[8
]}

Nitzsche, Anja ^{[2
]}

Hegemann, Bjoern

Bird, Alexander W. ^{[2
]}

Pelletier, Laurence ^{[2
,5
]}

Kittler, Ralf ^{[2
,6
,7
]}

Hua, Sujun ^{[6
,7
]}

Naumann, Ronald ^{[2
]}

Augsburg, Martina ^{[2
]}

Sykora, Martina M. ^{[3
]}

Hofemeister, Helmut ^{[1
]}

Zhang, Youming ^{[9
]}

Nasmyth, Kim ^{[10
]}

White, Kevin P. ^{[6
,7
]}

Dietzel, Steffen ^{[8
]}

Mechtler, Karl ^{[3
]}

Durbin, Richard ^{[4
]}

Stewart, A. Francis ^{[1
]}

Peters, Jan-Michael ^{[3
]}

Buchholz, Frank ^{[2
]}

Hyman, Anthony A. ^{[2
]}

机构：

[1] Tech Univ Dresden, Bioinnovat Ctr, D-01307 Dresden, Germany

[2] Max Planck Inst Mol Cell Biol & Genet, D-01307 Dresden, Germany

[3] Res Inst Mol Pathol, A-1030 Vienna, Austria

[4] Wellcome Trust Sanger Inst, Cambridge CB10 1HH, England

[5] Mt Sinai Hosp, Samuel Lunenfeld Res Inst, Toronto, ON M5G 1X5, Canada

[6] Univ Chicago, Inst Genom & Syst Biol, Chicago, IL 60637 USA

[7] Univ Chicago, Dept Human Genet, Chicago, IL 60637 USA

[8] Univ Munich, Munich, Germany

[9] Gene Bridges GmbH, Bioinnovationszentrum, Dresden, Germany

[10] Univ Oxford, Dept Biochem, Oxford OX1 3QU, England

来源：

NATURE METHODS | 2008年 / 5卷 / 05期

基金：

英国惠康基金;

关键词：

D O I：

10.1038/NMETH.1199

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

The interpretation of genome sequences requires reliable and standardized methods to assess protein function at high throughput. Here we describe a fast and reliable pipeline to study protein function in mammalian cells based on protein tagging in bacterial artificial chromosomes ( BACs). The large size of the BAC transgenes ensures the presence of most, if not all, regulatory elements and results in expression that closely matches that of the endogenous gene. We show that BAC transgenes can be rapidly and reliably generated using 96-well-format recombineering. After stable transfection of these transgenes into human tissue culture cells or mouse embryonic stem cells, the localization, protein-protein and/or protein-DNA interactions of the tagged protein are studied using generic, tag-based assays. The same high-throughput approach will be generally applicable to other model systems.

引用

页码：409 / 415

页数：7

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