Biology subjects

Marken, J. P.

Publications and source records attributed to Marken, J. P..

10 recordsLinked to original sources

bioRxiv2026

Flux-species sampling enables holistic exploration of biocircuit behavior

Biocircuit behavior is usually discovered by drawing uncertain kinetic parameters and simulating, but the scored phenotypes are functions of steady-state species, catalytic fluxes, dominance relations, and local response matrices. We introduce flux-species sampling (FSS), which samples that functional layer directly and algebraically inverts to parameters. This coordinate change turns regulatory modes into simple dominance chambers, gives every full-dimensional behavior-realizing regime positive measure, and can replace the sampling stage of existing pipelines without changing downstream stability or phenotype tests. A sequestration toy shows how a large species regime can collapse into a narrow parameter strip. In an enzymatic feedback motif, FSS rescues free-A* adaptation missed by parameter sampling and identifies the regime as non-Michaelian and complex-dominated. In MultiFate-N, FSS separates a sampling limitation from a structural limitation and rescues parameter sets realizing all 2N states. HighlightsO_LIFlux-species sampling (FSS) draws the control/function layer where biocircuit behavior is read C_LIO_LIFSS boxes intersect every full-dimensional behavior-realizing regime with positive measure C_LIO_LIFSS is no slower than parameter-space sampling and is a hot fix for existing pipelines C_LIO_LIFSS rescues non-Michaelian free-A* adaptation missed by parameter sampling C_LIO_LIFSS rescues the all-2N symmetric MultiFate regime missed by fixed parameter boxes C_LI eTOC blurbThis study reframes biocircuit exploration by sampling steady-state species and fluxes rather than raw parameters. The resulting flux-species coordinate exposes regulatory dominance regimes, rescues adaptation and multistability regimes missed by parameter sampling, and gives finite sampling nulls an explicit scope.

systems biology↗

bioRxiv2026

Sentinel Plants Enable Aboveground Detection of Belowground Soil Microbial Activity

Rhizosphere microbial processes play a central role in soil function and plant health yet remain difficult to monitor noninvasively. Engineered sentinel plants that use bacterial-to-plant communication channels are promising. However, no such efforts have thus far enabled a detectable aboveground response in the sentinel plant. Here, we optimize a previously described synthetic bacteria-to-plant communication channel based on the p-coumaroyl-homoserine lactone (pC-HSL) signaling molecule in plants to function as aboveground sentinels of belowground microbial activities. Arabidopsis thaliana sentinel plants harboring this optimized circuit detect root-applied pC-HSL at concentrations as low as 30 nM in roots and 3 M in leaves, demonstrating long-distance signal transmission from below ground to aboveground tissues. Moreover, sentinel plants report pC-HSL production by engineered Escherichia coli and Pseudomonas putida colonizing plant roots in both plate and soil assays. These results establish an engineered plant platform that converts rhizosphere microbial activity into a visible aboveground signal, enabling a minimally invasive platform for monitoring rhizosphere microbial gene expression and for precision agriculture and soil management.

bioengineering↗

bioRxiv2026

Quantitative measurement of synthetic repression curves reveals design challenges for genetic circuit engineering under growth arrest

Despite the fact that microbes in natural environments spend most of their time in growth arrest, we understand little about how this physiological state affects the performance of engineered genetic circuits. Here, we measure repression curves from a library of genetic NOT gates at single-cell resolution in Escherichia coli under both active growth and growth arrest to systematically investigate how growth arrest affects circuit behavior. We find that the impact of growth arrest on circuit performance is almost entirely dominated by a single effect: a >100-fold reduction in unrepressed expression levels. Growth arrest caused gene expression noise to increase moderately and had only minimal impacts on the sensitivity and sharpness of the repression curves. Our work shows both that conventional genetic circuit design paradigms are currently insufficient to develop circuits that can function properly under growth arrest, but also that addressing the reduction in just a single performance parameter would be sufficient to resolve this problem. This work expands our understanding of bacterial gene regulation under growth arrest and lays the groundwork for new design paradigms that will be essential in ensuring the safe and reliable performance of synthetic biology systems in real-world environments. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/703179v1_ufig1.gif" ALT="Figure 1"> View larger version (14K): org.highwire.dtl.DTLVardef@104ecd2org.highwire.dtl.DTLVardef@57f992org.highwire.dtl.DTLVardef@1c215c2org.highwire.dtl.DTLVardef@122a8c5_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2025

Association of E. coli Nissle with insect-parasitic nematode Steinernema hermaphroditum

Steinernema species are soil-dwelling and insect-parasitic nematodes that associate with symbiotic Xenorhabdus bacteria. During the infective juvenile (IJ) stage, Steinernema nematodes package species-specific Xenorhabdus bacteria in the anterior intestinal pockets. The nematodes can survive in the soil for months as they seek insect prey. The mechanisms of how these nematodes associate with environmental microbes other than their Xenorhabdus symbionts is barely known. Here, we report a new mechanism of E. coli Nissle (EcN) association with the nematode Steinernema hermaphroditum. We show that EcN cells are enclosed and lysed in at least four pairs of coelomocytes, suggesting these immune cells respond to bacterial invasion. During the IJ stage of nematode development, EcN cells localize to posterior intestinal vacuoles and enter the inter-cuticular space, where they proliferate, aggregate, then lyse. EcN cells expressed proteins in the cell lysates were maintained in the nematode cuticle over eight weeks in non-sterile soil. These observations suggest sequential steps of EcN colonization in the host nematodes involving an immune response that is distinctive from interactions with mutualistic symbiont. Our work establishes a novel framework of nematode-bacteria interaction with potential applications in environmental bioengineering.

microbiology↗

bioRxiv2022

Addressable and adaptable intercellular communication via DNA messaging

Engineered consortia are a major research focus for synthetic biologists because they can implement sophisticated behaviors inaccessible to single-strain systems. However, this functional capacity is constrained by their constituent strains ability to engage in complex communication. DNA messaging, by enabling information-rich channel-decoupled communication, is a promising candidate architecture for implementing complex communication. But its major advantage, its messages dynamic mutability, is still unexplored. We develop a framework for addressable and adaptable DNA messaging that leverages all three of these advantages and implement it in a plasmid conjugation-based communication channel. Our system can bias the transfer of messages to targeted receiver strains by 100-to 1000-fold, and their recipient lists can be dynamically updated in situ to control the flow of information through the population. This work lays the foundation for future developments that further utilize the unique advantages of DNA messaging to engineer previously-inaccessible levels of complexity into biological systems.

synthetic biology↗

bioRxiv2020

A geometric and structural approach to the analysis and design of biological circuit dynamics: a theory tailored for synthetic biology

Much of the progress in developing our ability to successfully design genetic circuits with predictable dynamics has followed the strategy of molding biological systems to fit into conceptual frameworks used in other disciplines, most notably the engineering sciences. Because biological systems have fundamental differences from systems in these other disciplines, this approach is challenging and the insights obtained from such analyses are often not framed in a biologically-intuitive way. Here, we present a new theoretical framework for analyzing the dynamics of genetic circuits that is tailored towards the unique properties associated with biological systems and experiments. Our framework approximates a complex circuit as a set of simpler circuits, which the system can transition between by saturating its various internal components. These approximations are connected to the intrinsic structure of the system, so this representation allows the analysis of dynamics which emerge solely from the systems structure. Using our framework, we analyze the presence of structural bistability in a leaky autoactivation motif and the presence of structural oscillations in the Repressilator.

synthetic biology↗

bioRxiv2019

Robust Estimation of Bacterial Cell Count from Optical Density

Optical density (OD) is a fast, cheap, and high-throughput measurement widely used to estimate the density of cells in liquid culture. These measurements, however, cannot be compared between instruments without a standardized calibration protocol and are challenging to relate to actual cell count. We address these shortcomings with an interlaboratory study comparing three OD calibration protocols, as applied to eight strains of E. coli engineered to constitutively express varying levels of GFP. These three protocols--comparison with colloidal silica (LUDOX), serial dilution of silica microspheres, and a reference colony-forming unit (CFU) assay--are all simple, low-cost, and highly accessible. Based on the results produced by the 244 teams completing this interlaboratory study, we recommend calibrating OD using serial dilution of silica microspheres, which readily produces highly precise calibration (95.5% of teams having residuals less than 1.2-fold), is easily assessed for quality control, and as a side effect also assesses the effective linear range of an instrument. Moreover, estimates of cell count from silica microspheres can be combined with fluorescence calibration against fluorescein to obtain units of Molecules of Equivalent Fluorescein (MEFL), allowing direct comparison and data fusion with equivalently calibrated flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.

synthetic biology↗

bioRxiv2019

Addressable, \"Packet-Based\" Intercellular Communication through Plasmid Conjugation

We develop a system for implementing \"packet-based\" intercellular communication in an engineered bacterial population via conjugation. Our system uses gRNA-based identification markers that allow messages to be addressed to specific strains via Cas9-mediated cleavage of messages sent to the wrong recipient, which we show reduces plasmid transfer by four orders of magnitude. Integrase-mediated editing of the address on the message plasmid allows cells to dynamically update the messages recipients in vivo. As a proof-of-concept demonstration of our system, we propose a linear path scheme that would propagate a message sequentially through the strains of a population in a defined order.

synthetic biology↗

bioRxiv2018

Modular, part-based control of gene expression response time using protein degradation tags

A fundamental principle of cellular signal processing is the encoding of information within the temporal dynamics of regulatory circuits. If synthetic circuits are to achieve the versatility and effectiveness of naturally-occurring circuits, it is necessary to develop simple, effective methods for the control of the dynamical properties of genetic circuits. However, current approaches to dynamical control often require extensive rewiring of circuit architecture, which hinders their implementation in a variety of systems. Therefore, it is essential that simple, modular, genetic parts-based frameworks are created to control the dynamical properties of circuits. Here we address this need by implementing a modular, genetic parts-based system which tunes the response time of a genes expression by tuning its degradation rate via the application of protein degradation tags with various affinities to their protease. This system provides a simple, easily- applicable framework for controlling the temporal aspects of genetic circuit behavior.

bioengineering↗

bioRxiv2018

The Genetic Insulator RiboJ Increases Expression of Insulated Genes

The self-cleaving ribozyme RiboJ is an insulator commonly used in genetic circuits to prevent unexpected interactions between neighboring parts. These interactions can compromise the modularity of the circuit, impeding the implementation of predictable genetic constructs. Despite its utility as an insulator, a quantitative assessment of the effect of RiboJ on the properties of downstream genetic parts is lacking. Here, we characterized the impact of insulation with RiboJ on expression of a reporter gene driven by a promoter from a library of 24 frequently employed constitutive promoters. We show that depending on the strength of the promoters, insulation with RiboJ increased protein abundance between twofold and tenfold and increased transcript abundance by an average of twofold. This result is the first to demonstrate that genetic insulators can impact the expression of downstream genes, potentially hindering the design of predictable genetic circuits and constructs.

synthetic biology↗