A smart polymer for sequence-selective binding, pulldown, and release of DNA targets

Elisha Krieg,William M Shih,Susanne Boye,Andreas Dahl,Albena Lederer,Krishna Gupta,Mathias Lesche

doi:10.1038/s42003-020-1082-2

Abstract

Selective isolation of DNA is crucial for applications in biology, bionanotechnology, clinical diagnostics and forensics. We herein report a smart methanol-responsive polymer (MeRPy) that can be programmed to bind and separate single- as well as double-stranded DNA targets. Captured targets are quickly isolated and released back into solution by denaturation (sequence-agnostic) or toehold-mediated strand displacement (sequence-selective). The latter mode allows 99.8% efficient removal of unwanted sequences and 79% recovery of highly pure target sequences. We applied MeRPy for the depletion of insulin, glucagon, and transthyretin cDNA from clinical next-generation sequencing (NGS) libraries. This step improved the data quality for low-abundance transcripts in expression profiles of pancreatic tissues. Its low cost, scalability, high stability and ease of use make MeRPy suitable for diverse applications in research and clinical laboratories, including enhancement of NGS libraries, extraction of DNA from biological samples, preparative-scale DNA isolations, and sorting of DNA-labeled non-nucleic acid targets.

Highlights

Selective isolation of DNA is crucial for applications in biology, bionanotechnology, clinical diagnostics and forensics
We show that methanol-responsive polymer (MeRPy) can bind one or multiple DNA targets, isolate, and release selected targets back into the medium
In a first practical example of this method we show that MeRPy is uniquely applicable for the enhancement of gene expression profile data obtained via RNA sequencing (RNA-seq)

Summary

Introduction

Selective isolation of DNA is crucial for applications in biology, bionanotechnology, clinical diagnostics and forensics. Captured targets are quickly isolated and released back into solution by denaturation (sequence-agnostic) or toehold-mediated strand displacement (sequence-selective). The latter mode allows 99.8% efficient removal of unwanted sequences and 79% recovery of highly pure target sequences. Scalability, high stability and ease of use make MeRPy suitable for diverse applications in research and clinical laboratories, including enhancement of NGS libraries, extraction of DNA from biological samples, preparative-scale DNA isolations, and sorting of DNA-labeled non-nucleic acid targets. It is crucial to engineer customizable, robust, and inexpensive materials that allow fast and efficient binding, manipulation, and release of selectable target sequences To address this challenge, we have developed an oligonucleotide-grafted methanol-responsive polymer (MeRPy). The pellet is redispersed in water and targets are released, either non-selectively by thermal or basic denaturation, or selectively via toehold-mediated strand displacement[16] (TMSD)

Methods

Results

Conclusion