Title: Design and Characterization of DNA Strand-Displacement Circuits in Serum-Supplemented Cell Medium

Abstract

The functional stability and lifetimes of synthetic molecular circuits in biological environments are important for long-term, stable sensors or controllers of cell or tissue behavior. DNA-based molecular circuits, particularly DNA strand-displacement circuits, provide simple and effective biocompatible control mechanisms and sensors, but are vulnerable to digestion by nucleases present in living tissues and serum-supplemented cell culture. The stability of double-stranded and single-stranded DNA circuit components in serum-supplemented cell medium and the corresponding effect of nuclease-mediated degradation on circuit performance were characterized to determine the major routes of degradation and DNA strand-displacement circuit failure. Simple circuit design choices, such as the use of 5' toeholds within the DNA complexes used as reactants in the strand-displacement reactions and the termination of single-stranded components with DNA hairpin domains at the 3' termini, significantly increase the functional lifetime of the circuit components in the presence of nucleases. Furthermore, simulations of multireaction circuits, guided by the experimentally measured operation of single-reaction circuits, enable predictive realization of multilayer and competitive-reaction circuit behavior. Altogether, these results provide a basic route to increased DNA circuit stability in cell culture environments.

Authors:

Fern, Joshua  ^[1];

• Search DOE PAGES for author "Fern, Joshua"
• Search DOE PAGES for ORCID "0000-0002-5028-2793"

Schulman, Rebecca  ^[2]

• Search DOE PAGES for author "Schulman, Rebecca"
• Search DOE PAGES for ORCID "0000-0003-4555-3162"

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+ Show Author Affiliations

1. Johns Hopkins Univ., Baltimore, MD (United States). Chemical and Biomolecular Engineering
2. Johns Hopkins Univ., Baltimore, MD (United States). Chemical and Biomolecular Engineering and Computer Science

Publication Date:

Tue May 30 00:00:00 EDT 2017

Research Org.:

Johns Hopkins Univ., Baltimore, MD (United States)

Sponsoring Org.:

USDOE Office of Science (SC), Basic Energy Sciences (BES)

OSTI Identifier:

1363867

Grant/Contract Number:  

SC0015906

Resource Type:

Accepted Manuscript

Journal Name:

ACS Synthetic Biology

Additional Journal Information:

Journal Volume: 6; Journal Issue: 9; Journal ID: ISSN 2161-5063

Publisher:

American Chemical Society (ACS)

Country of Publication:

United States

Language:

English

Subject:

77 NANOSCIENCE AND NANOTECHNOLOGY; 59 BASIC BIOLOGICAL SCIENCES; degradation; DNA strand displacement; molecular circuits; nuclease; serum