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- Title
High-throughput, genome-scale protein production method based on the wheat germ cell-free expression system.
- Authors
Endo, Yaeta; Sawasaki, Tatsuya
- Abstract
Current cell-free protein expression systems are capable of synthesizing proteins with high speed and accuracy; however, the yields are low due to their instability over time. Escherichia coli based systems are not always sufficient for expression of eukaryotic proteins. This report reviews a high-throughput protein production method based on the cell-free system prepared from eukaryote, wheat embryos. We first demonstrate a method for preparation of this extract that exhibited a high degree of stability and activity. To maximize translation yield and throughput, we address and resolve the following issues: (1) optimization of the ORF flanking regions; (2) PCR-based generation of DNA for mRNA production; (3) expression vectors for large-scale protein production; and (4) a translation reaction that does not require a membrane. The combination of these elemental processes with robotic automation resulted in high-throughput protein synthesis.
- Publication
Journal of structural and functional genomics, 2004, Vol 5, Issue 1-2, p45
- ISSN
1345-711X
- Publication type
Journal Article
- DOI
10.1023/B:JSFG.0000029208.83739.49