title: A chemically synthesized peptoid-based drag-tag enhances free-solution DNA sequencing by capillary electrophoresis.
abstract: 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.
authors.affiliation: Department of Chemistry, Northwestern University, 2145 N. Sheridan Road, Evanston, IL 60208, USA.
authors.forename: Russell D
authors.initials: RD
authors.lastname: Haynes
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Meagher
authors.forename: Annelise E
authors.initials: AE
authors.lastname: Barron
mesh_terms: D004247:DNA / Q000737:chemistry*; D019075:Electrophoresis, Capillary / Q000379:methods; D005998:Glycine / Q000737:chemistry*; D015394:Molecular Structure; D034444:Peptoids / Q000138:chemical synthesis* / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry; D017422:Sequence Analysis, DNA; D012996:Solutions
chemical_list: D034444:Peptoids; D012996:Solutions; D011092:Polyethylene Glycols; D004247:DNA; D005998:Glycine