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Chembiochem. 2016 Sep 2;17(17):1628-35. doi: 10.1002/cbic.201600298. Epub 2016 Jul 6.

Peptide Synthesis on a Next-Generation DNA Sequencing Platform.

Author information

1
Department of Pharmacology, Weill Cornell Medical College, Cornell University, New York, NY, 10065, USA.
2
Department of Biochemistry, Colorado State University, Fort Collins, CO, 80523, USA.
3
Department of Pharmacology, Weill Cornell Medical College, Cornell University, New York, NY, 10065, USA. srj2003@med.cornell.edu.

Abstract

Methods for displaying large numbers of peptides on solid surfaces are essential for high-throughput characterization of peptide function and binding properties. Here we describe a method for converting the >10(7) flow cell-bound clusters of identical DNA strands generated by the Illumina DNA sequencing technology into clusters of complementary RNA, and subsequently peptide clusters. We modified the flow-cell-bound primers with ribonucleotides thus enabling them to be used by poliovirus polymerase 3D(pol) . The primers hybridize to the clustered DNA thus leading to RNA clusters. The RNAs fold into functional protein- or small molecule-binding aptamers. We used the mRNA-display approach to synthesize flow-cell-tethered peptides from these RNA clusters. The peptides showed selective binding to cognate antibodies. The methods described here provide an approach for using DNA clusters to template peptide synthesis on an Illumina flow cell, thus providing new opportunities for massively parallel peptide-based assays.

KEYWORDS:

aptamers; high-throughput screening; mRNA display; next-generation DNA sequencing; peptide display; primer-dependent RNA synthesis

PMID:
27385640
PMCID:
PMC5183537
DOI:
10.1002/cbic.201600298
[Indexed for MEDLINE]
Free PMC Article

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