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The advent of easy-to-use open source microcontrollers, off-the-shelf electronics and customizable manufacturing technologies has facilitated the development of inexpensive scientific devices and laboratory equipment. In this study, we describe an imaging system that integrates low-cost and open-source hardware, software and genetic resources. The multi-fluorescence imaging system consists of readily available 470 nm LEDs, a Raspberry Pi camera and a set of filters made with low cost acrylics. This device allows imaging in scales ranging from single colonies to entire plates. We developed a set of genetic components (e.g. promoters, coding sequences, terminators) and vectors following the standard framework of Golden Gate, which allowed the fabrication of genetic constructs in a combinatorial, low cost and robust manner. In order to provide simultaneous imaging of multiple wavelength signals, we screened a series of long stokes shift fluorescent proteins that could be combined with cyan/green fluorescent proteins. We found CyOFP1, mBeRFP and sfGFP to be the most compatible set for 3-channel fluorescent imaging. We developed open source Python code to operate the hardware to run time-lapse experiments with automated control of illumination and camera and a Python module to analyze data and extract meaningful biological information. To demonstrate the potential application of this integral system, we tested its performance on a diverse range of imaging assays often used in disciplines such as microbial ecology, microbiology and synthetic biology. We also assessed its potential use in a high school environment to teach biology, hardware design, optics, and programming. Together, these results demonstrate the successful integration of open source hardware, software, genetic resources and customizable manufacturing to obtain a powerful, low cost and robust system for education, scientific research and bioengineering. All the resources developed here are available under open source licenses.

Access to affordable and reliable scientific instrumentation remains a significant barrier to the democratization of healthcare and scientific research. In the field of biotechnology, in particular, the complexity, cost, and infrastructure requirements of many instruments continue to limit their accessibility, especially in resource-limited environments. Despite the recent increase in the development of open-source tools, driven by advances in digital fabrication and electronic prototyping, few of these projects have reached large-scale implementation or validation in real-world settings. Here, we present qByte, an open-source, 8-tube isothermal fluorimeter designed to overcome these barriers by offering a cost-effective ($60) yet production-ready solution. qByte leverages standard digital manufacturing and Printed Circuit Board (PCB) assembly techniques and is designed to be portable, making it ideal for both laboratory and field use. The device has been benchmarked against commercial real-time thermocyclers and spectrophotometers, showing comparable results across four key applications: nucleic acid amplification and detection, including the on-site diagnosis of human parasites in Ghana, analysis of protein activity and stability, genetic construct characterization, and bacterial viability tests. Taken together, our results proved qByte as flexible and reliable equipment for a variety of biological tests and applications, while its affordability and open-source design simplify further development and allow adaptation to the needs of future users.

The urgent need to develop a more equitable bioeconomy has positioned biotechnology capacity building at the forefront of international priorities. However, in many educational institutions, particularly in low- and middle-income countries, this remains a major challenge due to limited access to reagents, equipment, and technical documentation. In this work, we describe Open Educational Resources (OER) composed of locally produced biological reagents, open source hardware and free software to teach fundamental techniques in biotechnology such as LAMP DNA amplification, RT-PCR RNA detection, enzyme kinetics and fluorescence imaging. The use of locally produced reagents and devices reduces costs by up to one order of magnitude. During the pandemic lockdowns, these tools were distributed nationwide to students' homes as a lab-in-a-box for remote teaching of molecular biology. To test their performance, a total of 93 undergraduate students tested these resources during a biochemistry practical course. 27 out of 31 groups (~87%) successfully achieved the objectives of the PCR activity, while 28 out of 31 groups (~90%) correctly identified the target using LAMP reactions. To assess the potential application in secondary school, we organized three workshops for high school teachers from different institutions across Chile and performed an anonymous questionnaire, obtaining a strong agreement on how these OER broaden teachers' perspectives on the techniques and facilitate the teaching of molecular biology topics. This effort was made possible through a close collaboration with open source technology advocates and members of DIYbio communities, whose work is paving the way for low-cost training in biology. All the protocols and design files are available under open source licenses.

The development of effective tools to combat viral diseases remains a major challenge for the aquaculture industry. Infectious pancreatic necrosis virus (IPNv) is one of the most devastating pathogens affecting salmonids, leading to high mortality rates and substantial economic losses worldwide. Here, we present a novel nanobody discovery pipeline based on a Type IIS restriction enzyme-driven library assembly method that enables the rapid generation of highly diverse nanobody repertoires. This streamlined approach not only shortens the time required for nanobody identification but also offers remarkable adaptability, allowing its application to virtually any protein target, including antigens from aquaculture pathogens and beyond. By integrating this strategy with density gradient–based enrichment and high-throughput screening, we successfully identified and validated a nanobody against the VP2 protein of IPNv, a key structural component essential for viral infectivity. These findings highlight the potential of this platform both as a versatile methodological advance in antibody engineering and as a practical foundation for developing innovative diagnostic and therapeutic tools. Ultimately, nanobodies generated through this pipeline could play a pivotal role in improving disease management and enhancing sustainability in aquaculture.

The urgent need to develop a more equitable bioeconomy has positioned biotechnology capacity building at the forefront of international priorities. However, in many educational institutions, particularly in low- and middle-income countries, this remains a major challenge due to limited access to reagents, equipment, and technical documentation. In this work, we describe Open Educational Resources (OER) composed of locally produced biological reagents, open source hardware and free software to teach fundamental techniques in biotechnology such as LAMP DNA amplification, RT-PCR RNA detection, enzyme kinetics and fluorescence imaging. The use of locally produced reagents and devices reduces costs by up to one order of magnitude. During the pandemic lockdowns, these tools were distributed nationwide to students’ homes as a lab-in-a-box for remote teaching of molecular biology. To test their performance, a total of 93 undergraduate students tested these resources during a biochemistry practical course. 27 out of 31 groups (~87%) successfully achieved the objectives of the PCR activity, while 28 out of 31 groups (~90%) correctly identified the target using LAMP reactions. To assess the potential application in secondary school, we organized three workshops for high school teachers from different institutions across Chile and performed an anonymous questionnaire, obtaining a strong agreement on how these OER broaden teachers’ perspectives on the techniques and facilitate the teaching of molecular biology topics. This effort was made possible through a close collaboration with open source technology advocates and members of DIYbio communities, whose work is paving the way for low-cost training in biology. All the protocols and design files are available under open source licenses.

Bidirectional intercellular signaling is an essential feature of multicellular organisms, and the engineering of complex biological systems will require multiple pathways for intercellular signaling with minimal crosstalk. Natural quorum‐sensing systems provide components for cell communication, but their use is often constrained by signal crosstalk. We have established new orthogonal systems for cell–cell communication using acyl homoserine lactone signaling systems. Quantitative measurements in contexts of differing receiver protein expression allowed us to separate different types of crosstalk between 3‐oxo‐C6‐ and 3‐oxo‐C12‐homoserine lactones, cognate receiver proteins, and DNA promoters. Mutating promoter sequences minimized interactions with heterologous receiver proteins. We used experimental data to parameterize a computational model for signal crosstalk and to estimate the effect of receiver protein levels on signal crosstalk. We used this model to predict optimal expression levels for receiver proteins, to create an effective two‐channel cell communication device. Establishment of a novel spatial assay allowed measurement of interactions between geometrically constrained cell populations via these diffusible signals. We built relay devices capable of long‐range signal propagation mediated by cycles of signal induction, communication and response by discrete cell populations. This work demonstrates the ability to systematically reduce crosstalk within intercellular signaling systems and to use these systems to engineer complex spatiotemporal patterning in cell populations. Synopsis The use of multiple homoserine lactone quorum‐sensing signals in synthetic circuits has been hampered by crosstalk. Measurement and modeling of the system allows for rational modifications to minimize crosstalk and create programmed spatial behavior. Quantitative measurements were used to build and parameterize a cellular model of synthetic signal transduction. Rationally designed changes to promoter sequences combined with model‐informed tuning of receiver protein expression reduce crosstalk to undetectable levels. A novel spatial assay allows measurement of confined bacterial populations in defined geometries. Relay devices demonstrate bidirectional communication resulting in initiation and spatial propagation of a signal. Graphical Abstract The use of multiple homoserine lactone quorum‐sensing signals in synthetic circuits has been hampered by crosstalk. Measurement and modeling of the system allows for rational modifications to minimize crosstalk and create programmed spatial behavior.

Background Understanding how spatial patterns of gene expression emerge from the interaction of individual gene networks is a fundamental challenge in biology. Developing a synthetic experimental system with a common theoretical framework that captures the emergence of short- and long-range spatial correlations (and anti-correlations) from interacting gene networks could serve to uncover generic scaling properties of these ubiquitous phenomena. Results Here, we combine synthetic biology, statistical mechanics models, and computational simulations to study the spatial behavior of synthetic gene networks (SGNs) in Escherichia coli quasi-2D colonies growing on hard agar surfaces. Guided by the combined mechanisms of the contact process lattice simulation and two-dimensional Ising model (CPIM), we describe the spatial behavior of bi-stable and chemically coupled SGNs that self-organize into patterns of long-range correlations with power-law scaling or short-range anti-correlations. These patterns, resembling ferromagnetic and anti-ferromagnetic configurations of the Ising model near critical points, maintain their scaling properties upon changes in growth rate and cell shape. Conclusions Our findings shed light on the spatial biology of coupled and bistable gene networks in growing cell populations. This emergent spatial behavior could provide insights into the study and engineering of self-organizing gene patterns in eukaryotic tissues and bacterial consortia.

• High-efficiency methods for DNA assembly have enabled the routine assembly of synthetic DNAs of increased size and complexity. However, these techniques require customization, elaborate vector sets or serial manipulations for the different stages of assembly. • We have developed Loop assembly based on a recursive approach to DNA fabrication. The system makes use of two Type IIS restriction endonucleases and corresponding vector sets for efficient and parallel assembly of large DNA circuits. Standardized level 0 parts can be assembled into circuits containing 1, 4, 16 or more genes by looping between the two vector sets. • The vectors also contain modular sites for hybrid assembly using sequence overlap methods. - Loop assembly enables efficient and versatile DNA fabrication for plant transformation. We show the construction of plasmids up to 16 genes and 38 kb with high efficiency (> 80%). We have characterized Loop assembly on over 200 different DNA constructs and validated the fidelity of the method by high-throughput Illumina plasmid sequencing. • Our method provides a simple generalized solution for DNA construction with standardized parts. The cloning system is provided under an OpenMTA license for unrestricted sharing and open access.

Agrobacterium rhizogenes (or Rhizobium rhizogenes) is able to transform plant genomes and induce the production of hairy roots. We describe the use of A. rhizogenes in tomato (Solanum spp.) to rapidly assess gene expression and function. Gene expression of reporters is indistinguishable in plants transformed by Agrobacterium tumefaciens as compared with A. rhizogenes. A root cell type- and tissue-specific promoter resource has been generated for domesticated and wild tomato (Solanum lycopersicum and Solanum pennellii, respectively) using these approaches. Imaging of tomato roots using A. rhizogenes coupled with laser scanning confocal microscopy is facilitated by the use of a membrane-tagged protein fused to a red fluorescent protein marker present in binary vectors. Tomato-optimized isolation of nuclei tagged in specific cell types and translating ribosome affinity purification binary vectors were generated and used to monitor associated messenger RNA abundance or chromatin modification. Finally, transcriptional reporters, translational reporters, and clustered regularly interspaced short palindromic repeats-associated nuclease9 genome editing demonstrate that SHORT-ROOT and SCARECROW gene function is conserved between Arabidopsis (Arabidopsis thaliana) and tomato.

Automated cell segmentation in time-lapse confocal images of growing Arabidopsis combined with ratiometric imaging of fluorescent gene expression reporters permits the correlation of cellular properties with gene expression in the growing plant. We present the coupled use of specifically localized fluorescent gene markers and image processing for automated quantitative analysis of cell growth and genetic activity across living plant tissues. We used fluorescent protein markers to identify cells, create seeds and boundaries for the automatic segmentation of cell geometries and ratiometrically measure gene expression cell by cell in Arabidopsis thaliana .