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Poster Presentations
Day 4, June 25（Wed.）
Room P (Maesato East, Foyer, Ocean Wing)
- 4P-PM-29
High-Throughput Synthesis of Stable Isotope-Labeled Peptides Using Synthetic ssDNA Oligo Pools.
(RIKEN BDR)
oReiko Nakagawa, Keiko Masuda, Aya Sato, Yoshihiro Shimizu
Quantitative proteomics using stable isotope-labeled internal standard peptides is a sensitive and accurate method for biomarker discovery and quantitative biology. However, existing peptide synthesis methods are costly, labor-intensive, and typically target human sequences, limiting their broader application.
We previously developed a method called MS-QBiC (MS-based quantification by isotope-labeled cell-free products), which synthesizes stable isotope-labeled peptides using the PURE system, an E. coli-based cell-free protein synthesis platform. The original approach involved designing primers for each peptide and amplifying them via PCR to prepare DNA templates for the PURE system.
In this study, we aimed to streamline the process by using synthetic ssDNA oligonucleotide pools as DNA templates. This enabled the simultaneous synthesis of approximately 200 peptides in a single reaction. Additionally, we incorporated unique barcode sequences at both ends of the ssDNA oligonucleotides, enabling selective amplification of specific peptide sets using PCR.
By integrating this approach with our MS-QBiC technology, we can now rapidly and cost-effectively prepare stable isotope-labeled peptides, overcoming previous limitations on the number of synthesized peptides or target species. This method provides a more flexible and accessible solution for quantitative proteomics.