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A chemically synthesized peptoid‐based drag‐tag enhances free‐solution DNA sequencing by capillary electrophoresis
Author(s) -
Haynes Russell D.,
Meagher Robert J.,
Barron Annelise E.
Publication year - 2011
Publication title -
peptide science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/bip.21627
Subject(s) - peptoid , chemistry , capillary electrophoresis , dna , electrophoresis , chromatography , drag , capillary action , combinatorial chemistry , peptide , biochemistry , thermodynamics , physics
We report a capillary‐based DNA sequencing read length of 100 bases in 16 min using end‐labeled free‐solution conjugate electrophoresis (FSCE) with a monodisperse poly‐N‐substituted glycine (polypeptoid) as a synthetic drag‐tag. FSCE enabled rapid separation of single‐stranded (ss) DNA sequencing fragments with single‐base resolution without the need for a viscous DNA separation matrix. Protein‐based drag‐tags previously used for FSCE sequencing, for example, streptavidin, are heterogeneous in molar mass (polydisperse); the resultant band‐broadening can make it difficult to obtain the single‐base resolution necessary for DNA sequencing. In this study, we synthesized and HPLC‐purified a 70mer poly‐N‐(methoxyethyl)glycine (NMEG) drag‐tag with a molar mass of ∼11 kDa. The NMEG monomers that comprise this peptoid drag‐tag are interesting for bioanalytical applications, because the methoxyethyl side chain's chemical structure is reminiscent of the basic monomer unit of polyethylene glycol, a highly biocompatible commercially available polymer, which, however, is not available in monodisperse preparation at an ∼11 kDa molar mass. This is the first report of ssDNA separation and of four‐color, base‐by‐base DNA sequencing by FSCE through the use of a chemically synthesized drag‐tag. These results show that high‐molar mass, chemically synthesized drag‐tags based on the polyNMEG structure, if obtained in monodisperse preparation, would serve as ideal drag‐tags and could help FSCE reach the commercially relevant read lengths of 100 bases or more. © 2011 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 96:702–707, 2011.