title: Separation efficiency of free-solution conjugated electrophoresis with drag-tags incorporating a synthetic amino acid.
abstract: DNA sequencing or separation by conventional capillary electrophoresis with a polymer matrix has some inherent drawbacks, such as the expense of polymer matrix and limitations in sequencing read length. As DNA fragments have a linear charge-to-friction ratio in free solution, DNA fragments cannot be separated by size. However, size-based separation of DNA is possible in free-solution conjugate electrophoresis (FSCE) if a "drag-tag" is attached to DNA fragments because the tag breaks the linear charge-to-friction scaling. Although several previous studies have demonstrated the feasibility of DNA separation by free-solution conjugated electrophoresis, generation of a monodisperse drag-tag and identification of a strong, site-specific conjugation method between a DNA fragment and a drag-tag are challenges that still remain. In this study, we demonstrate an efficient FSCE method by conjugating a biologically synthesized elastin-like polypeptide (ELP) and green fluorescent protein (GFP) to DNA fragments. In addition, to produce strong and site-specific conjugation, a methionine residue in drag-tags is replaced with homopropargylglycine (Hpg), which can be conjugated specifically to a DNA fragment with an azide site.
authors.affiliation: Department of Chemical Engineering, Hongik University, Seoul, Korea.
authors.forename: Kyung-Ho
authors.initials: KH
authors.lastname: Seo
authors.affiliation: Department of Chemical Engineering, Hongik University, Seoul, Korea.|Material Research Center, SAMSUNG ELECTRONICS Co, Suwon, Korea.
authors.forename: Hun-Su
authors.initials: HS
authors.lastname: Chu
authors.affiliation: Department of Molecular Science and Technology, Ajou University, Suwon, Korea.
authors.forename: Tae Hyeon
authors.initials: TH
authors.lastname: Yoo
authors.affiliation: Department of Chemical Engineering, Pusan National University, Pusan, Korea.
authors.forename: Sun-Gu
authors.initials: SG
authors.lastname: Lee
authors.affiliation: Department of Chemical Engineering, Hongik University, Seoul, Korea.
authors.forename: Jong-In
authors.initials: JI
authors.lastname: Won
mesh_terms: D000596:Amino Acids / Q000235:genetics*; D004247:DNA / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D004549:Elastin / Q000235:genetics; D004586:Electrophoresis / Q000379:methods*; D015336:Molecular Probe Techniques; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods*
chemical_list: D000596:Amino Acids; D011993:Recombinant Fusion Proteins; D004247:DNA; D004549:Elastin
keywords: Drag-tag; ELP; FSCE; GFP; Synthetic amino acid