Comblike, monodisperse polypeptoid drag-tags for DNA separations by end-labeled free-solution electrophoresis (ELFSE)
- Authors
- Haynes, R.D.; Meagher, R.J.; Won, J.-I.; Bogdan, F.M.; Barron, A.E.
- Issue Date
- 2005
- Citation
- Bioconjugate Chemistry, v.16, no.4, pp.929 - 938
- Journal Title
- Bioconjugate Chemistry
- Volume
- 16
- Number
- 4
- Start Page
- 929
- End Page
- 938
- URI
- https://scholarworks.bwise.kr/hongik/handle/2020.sw.hongik/25678
- DOI
- 10.1021/bc0496915
- ISSN
- 1043-1802
- Abstract
- The development of innovative technologies designed to reduce the cost and increase the throughput of DNA separations continues to be important for large-scale sequencing and genotyping efforts. We report research aimed at the further development of a free-solution bioconjugate method of DNA size separation by capillary electrophoresis (CE), in particular, the determination of an optimal molecular architecture for polyamide-based drag-tags. We synthesized several branched poly(N-methoxyethyl glycine)s (poly(NMEG)s, a class of polypeptoids) as novel friction-generating entities for end-on attachment to DNA molecules. A 30-mer poly(NMEG) backbone, comprising five evenly spaced reactive ε-amino groups, was synthesized on solid phase, cleaved, and purified to monodispersity by RP-HPLC. Three different comblike derivatives of this backbone molecule were created by (1) acetylating the ε-amino groups or (2) appending small, monodisperse NMEG oligomers (a tetramer and an octamer). Grafting of the oligo(NMEG)s was done using solution-phase amide bond formation chemistry. Once purified to total monodispersity, the three different drag-tags were studied by free-solution electrophoresis to observe the effect of branching on their hydrodynamic drag or α and hence their ability to separate DNA. Drag was found to scale linearly with total molecular weight, regardless of branch length. The octamer-branched drag-tag-DNA conjugate was used to separate ssDNA products of 50, 75, 100, and 150 bases in length by free-solution CE in less than 10 min. Hence, the use of branched or comblike drag-tags is both a feasible and an effective way to achieve high frictional drag, allowing the high-resolution separation of relatively large DNA molecules by free-solution CE without the need to synthesize very long polymers. © 2005 American Chemical Society.
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