Competitive binding-based optical DNA mapping for fast identification of bacteria - multi-ligand transfer matrix theory and experimental applications on Escherichia coli
Artikel i vetenskaplig tidskrift, 2014

We demonstrate a single DNA molecule optical mapping assay able to resolve a specific Escherichia coli strain from other strains. The assay is based on competitive binding of the fluorescent dye YOYO-1 and the AT-specific antibiotic netropsin. The optical map is visualized by stretching the DNA molecules in nanofluidic channels. We optimize the experimental conditions to obtain reproducible barcodes containing as much information as possible. We implement a multi-ligand transfer matrix method for calculating theoretical barcodes from known DNA sequences. Our method extends previous theoretical approaches for competitive binding of two types of ligands to many types of ligands and introduces a recursive approach that allows long barcodes to be calculated with standard computer floating point formats. The identification of a specific E. coli strain (CCUG 10979) is based on mapping of 50-160 kilobasepair experimental DNA fragments onto the theoretical genome using the developed theory. Our identification protocol introduces two theoretical constructs: aP-value for a best experiment-theory match and an information score threshold. The developed methods provide a novel optical mapping toolbox for identification of bacterial species and strains. The protocol does not require cultivation of bacteria or DNA amplification, which allows for ultra-fast identification of bacterial pathogens.

Författare

A. Nilsson

Lunds universitet

Gustav Emilsson

Chalmers, Teknisk fysik, Bionanofotonik

Lena Nyberg

Chalmers, Kemi- och bioteknik, Fysikalisk kemi

C. Noble

Lunds universitet

Liselott Svensson-Stadler

Göteborgs universitet

Joachim Fritzsche

Chalmers, Teknisk fysik, Kemisk fysik

Edward R.B. Moore

Göteborgs universitet

J. O. Tegenfeldt

Lunds universitet

T. Ambjornsson

Lunds universitet

Fredrik Westerlund

Chalmers, Kemi- och bioteknik, Fysikalisk kemi

Nucleic Acids Research

0305-1048 (ISSN) 1362-4962 (eISSN)

Vol. 42 15 e118- e118

Styrkeområden

Nanovetenskap och nanoteknik

Livsvetenskaper och teknik

Ämneskategorier

Biokemi och molekylärbiologi

DOI

10.1093/nar/gku556