| RRC ID |
3316
|
| Author |
Kawahashi Y, Doi N, Oishi Y, Tsuda C, Takashima H, Baba T, Mori H, Ito T, Yanagawa H.
|
| Title |
High-throughput fluorescence labelling of full-length cDNA products based on a reconstituted translation system.
|
| Journal |
J Biochem
|
| Abstract |
Although recent advances in fluorescence-based technologies, such as protein microarrays, have made it possible to analyse more than 10,000 proteins at once, there is a bottleneck in the step of preparation of large numbers of fluorescently labelled proteins for the comprehensive analysis of protein-protein interactions. Here we describe two independent methods for high-throughput fluorescence-labelling of full-length cDNA products at their C-termini using a reconstituted translation system containing fluorescent puromycin. For the first method, release factor-free systems were used. For the second method, stop codons were excluded from cDNAs by using a common mismatch primer in mutagenic PCR. These methods yielded large numbers of labelled proteins from cDNA sets of various organisms, such as mouse, yeast and Escherichia coli.
|
| Volume |
141(1)
|
| Pages |
19-24
|
| Published |
2007-1-1
|
| DOI |
10.1093/jb/mvm003
|
| PII |
mvm003
|
| PMID |
17148548
|
| MeSH |
Carbocyanines / chemistry
Cell-Free System / metabolism
DNA, Complementary / chemistry*
Escherichia coli / metabolism
Escherichia coli Proteins / metabolism
Fluorescent Dyes* / chemistry
Multienzyme Complexes / metabolism
Proteasome Endopeptidase Complex
Protein Biosynthesis*
Proteomics / methods
Puromycin / analogs & derivatives*
Saccharomyces cerevisiae Proteins / metabolism
|
| IF |
2.476
|
| Times Cited |
4
|
|
WOS Category
|
BIOCHEMISTRY & MOLECULAR BIOLOGY
|
| Resource |
| Prokaryotes E. coli |
ASKA library |
