There is a growing demand for custom synthesis of short genes libraries coding for active peptides or regulatory RNAs. DNA microarrays can be manufactured by synthesizing oligonucleotides on solid substrate in a massively-parallel manner using a high-yield low cost chemistry. Oligonucleotides can be cleaved off the microarray surface and recovered as a pool. Our hypothesis is that we can use this technology to create custom libraries of long DNA oligonucleotides at a much reduced cost and increased complexity compared to current technologies. Our long term objectives are to implement a commercial service of affordable custom synthesis of long oligonucleotide libraries. These libraries are used as a research tools in many applications such as studies on gene silencing, protein-DNA interaction, epitope mapping or even antimicrobial peptides. There are no limits for applications than the imagination of scientists. The heath relatedness of the project resides in the facts that these applications lead to the discovery of new cellular mechanisms, diagnosis tools, drugs or even vaccines. The scope of the proposed project is 1) to demonstrate the feasibility of using an emulsion-based PCR to amplify oligonucleotide libraries;2) to investigate the possibility to synthesize libraries of oligonucleotide up to 150 mer in length and 3) to determine the synthesis error rate and type of sequence mutations present in these libraries. We will in particular test the effect of droplet size and number of templates per droplet on the PCR amplification of oligonucleotide template in an emulsion. We wil characterize the complexity of an amplified library by deep-sequencing a PCR product. The large amount of sequence information obtained will also permit an in depth characterization the type of errors occuring during massively-parallel long oligonucleotide synthesis.
The unprecedented availability of affordable custom libraries of long oligonucleotides will enable new experimentations in fields such as gene silencing, protein-DNA interaction, epitope mapping or even antimicrobial peptides. This technology will undoubtedly bolster the discovery of new cellular mechanisms, diagnosis tools, drugs or even vaccines, ultimately benefiting the society.
| Murgha, Yusuf; Beliveau, Brian; Semrau, Kassandra et al. (2015) Combined in vitro transcription and reverse transcription to amplify and label complex synthetic oligonucleotide probe libraries. Biotechniques 58:301-7 |
| Beliveau, Brian J; Joyce, Eric F; Apostolopoulos, Nicholas et al. (2012) Versatile design and synthesis platform for visualizing genomes with Oligopaint FISH probes. Proc Natl Acad Sci U S A 109:21301-6 |
