TY - JOUR
T1 - Novel fluorescence labeling and high-throughput assay technologies for in vitro analysis of protein interactions
AU - Doi, Nobuhide
AU - Takashima, Hideaki
AU - Kinjo, Masataka
AU - Sakata, Kyoko
AU - Kawahashi, Yuko
AU - Oishi, Yuko
AU - Oyama, Rieko
AU - Miyamoto-Sato, Etsuko
AU - Sawasaki, Tatsuya
AU - Endo, Yaeta
AU - Yanagawa, Hiroshi
PY - 2002
Y1 - 2002
N2 - We developed and tested a simple method for fluorescence labeling and interaction analysis of proteins based on a highly efficient in vitro translation system combined with high-throughput technologies such as microarrays and fluorescence cross-correlation spectroscopy (FCCS). By use of puromycin analogs linked to various fluorophores through a deoxycytidylic acid linker, a single fluorophore can be efficiently incorporated into a protein at the carboxyl terminus during in vitro translation. We confirmed that the resulting fluorescently labeled proteins are useful for probing protein-protein and protein-DNA interactions by means of pulldown assay, DNA microarrays, and FCCS in model experiments. These fluorescence assay systems can be easily extended to highly parallel analysis of protein interactions in studies of functional genomics.
AB - We developed and tested a simple method for fluorescence labeling and interaction analysis of proteins based on a highly efficient in vitro translation system combined with high-throughput technologies such as microarrays and fluorescence cross-correlation spectroscopy (FCCS). By use of puromycin analogs linked to various fluorophores through a deoxycytidylic acid linker, a single fluorophore can be efficiently incorporated into a protein at the carboxyl terminus during in vitro translation. We confirmed that the resulting fluorescently labeled proteins are useful for probing protein-protein and protein-DNA interactions by means of pulldown assay, DNA microarrays, and FCCS in model experiments. These fluorescence assay systems can be easily extended to highly parallel analysis of protein interactions in studies of functional genomics.
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U2 - 10.1101/gr.218802. Article published online before print in February 2002
DO - 10.1101/gr.218802. Article published online before print in February 2002
M3 - Article
C2 - 11875038
AN - SCOPUS:18344365585
SN - 1088-9051
VL - 12
SP - 487
EP - 492
JO - Genome Research
JF - Genome Research
IS - 3
ER -