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Multiplexed DNA Identification Using Site Specific dCas9 Barcodes and Nanopore Sensing.

Accepted version
Peer-reviewed

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Authors

Ermann, Niklas 
Gutierrez, Richard 
Graham, James 

Abstract

Decorating double-stranded DNA with dCas9 barcodes to identify characteristic short sequences provides an alternative to fully sequencing DNA samples for rapid and highly specific analysis of a DNA sample. Solid state nanopore sensors are especially promising for this type of single-molecule sensing because of the ability to analyze patterns in the ionic current signatures of DNA molecules. Here, we systematically demonstrate the use of highly specific dCas9 probes to create unique barcodes on the DNA that can be read out using nanopore sensors. Single dCas9 probes are targeted to various positions on DNA strands up to 48 kbp long and are effectively measured in high salt conditions typical of nanopore sensing. Multiple probes bound to the same DNA strand at characteristic target sequences create distinct barcodes of double and triple peaks. Finally, double and triple barcodes are used to simultaneously identify two different DNA targets in a background mixture of bacterial DNA. Our method forms the basis of a fast and versatile assay for multiplexed DNA sensing applications in complex samples.

Description

Keywords

Cas9, DNA barcode, biosensor, nanopore sensing, single-molecule detection, Biosensing Techniques, DNA, Electrochemical Techniques, Fluorescent Dyes, Nanopores

Journal Title

ACS Sens

Conference Name

Journal ISSN

2379-3694
2379-3694

Volume Title

4

Publisher

American Chemical Society (ACS)

Rights

All rights reserved
Sponsorship
European Research Council (647144)