Brenda Wang

All Publications

• Semiautomated Production of Cell-Free Biosensors. ACS synthetic biology Brown, D. M., Phillips, D. A., Garcia, D. C., Arce, A., Lucci, T., Davies, J. P., Mangini, J. T., Rhea, K. A., Bernhards, C. B., Biondo, J. R., Blum, S. M., Cole, S. D., Lee, J. A., Lee, M. S., McDonald, N. D., Wang, B., Perdue, D. L., Bower, X. S., Thavarajah, W., Karim, A. S., Lux, M. W., Jewett, M. C., Miklos, A. E., Lucks, J. B. 2025

  Abstract

  Cell-free synthetic biology biosensors have potential as effective in vitro diagnostic technologies for the detection of chemical compounds, such as toxins and human health biomarkers. They have several advantages over conventional laboratory-based diagnostic approaches, including the ability to be assembled, freeze-dried, distributed, and then used at the point of need. This makes them an attractive platform for cheap and rapid chemical detection across the globe. Though promising, a major challenge is scaling up biosensor manufacturing to meet the needs of their multiple uses. Currently, cell-free biosensor assembly during lab-scale development is mostly performed manually by the operator, leading to quality control and performance variability issues. Here we explore the use of liquid-handling robotics to manufacture cell-free biosensor reactions. We compare both manual and semiautomated reaction assembly approaches using the Opentrons OT-2 liquid handling platform on two different cell-free gene expression assay systems that constitutively produce colorimetric (LacZ) or fluorescent (GFP) signals. We test the designed protocol by constructing an entire 384-well plate of fluoride-sensing cell-free biosensors and demonstrate that they perform close to expected detection outcomes.

  View details for DOI 10.1021/acssynbio.4c00703

  View details for PubMedID 40073441

• An Automated Cell-Free Workflow for Transcription Factor Engineering. ACS synthetic biology Ekas, H. M., Wang, B., Silverman, A. D., Lucks, J. B., Karim, A. S., Jewett, M. C. 2024

  Abstract

  The design and optimization of metabolic pathways, genetic systems, and engineered proteins rely on high-throughput assays to streamline design-build-test-learn cycles. However, assay development is a time-consuming and laborious process. Here, we create a generalizable approach for the tailored optimization of automated cell-free gene expression (CFE)-based workflows, which offers distinct advantages over in vivo assays in reaction flexibility, control, and time to data. Centered around designing highly accurate and precise transfers on the Echo Acoustic Liquid Handler, we introduce pilot assays and validation strategies for each stage of protocol development. We then demonstrate the efficacy of our platform by engineering transcription factor-based biosensors. As a model, we rapidly generate and assay libraries of 127 MerR and 134 CadR transcription factor variants in 3682 unique CFE reactions in less than 48 h to improve limit of detection, selectivity, and dynamic range for mercury and cadmium detection. This was achieved by assessing a panel of ligand conditions for sensitivity (to 0.1, 1, 10 muM Hg and 0, 1, 10, 100 muM Cd for MerR and CadR, respectively) and selectivity (against Ag, As, Cd, Co, Cu, Hg, Ni, Pb, and Zn). We anticipate that our Echo-based, cell-free approach can be used to accelerate multiple design workflows in synthetic biology.

  View details for DOI 10.1021/acssynbio.4c00471

  View details for PubMedID 39373325

• Engineering a PbrR-Based Biosensor for Cell-Free Detection of Lead at the Legal Limit. ACS synthetic biology Ekas, H. M., Wang, B., Silverman, A. D., Lucks, J. B., Karim, A. S., Jewett, M. C. 2024

  Abstract

  Industrialization and failing infrastructure have led to a growing number of irreversible health conditions resulting from chronic lead exposure. While state-of-the-art analytical chemistry methods provide accurate and sensitive detection of lead, they are too slow, expensive, and centralized to be accessible to many. Cell-free biosensors based on allosteric transcription factors (aTFs) can address the need for accessible, on-demand lead detection at the point of use. However, known aTFs, such as PbrR, are unable to detect lead at concentrations regulated by the Environmental Protection Agency (24-72 nM). Here, we develop a rapid cell-free platform for engineering aTF biosensors with improved sensitivity, selectivity, and dynamic range characteristics. We apply this platform to engineer PbrR mutants for a shift in limit of detection from 10 μM to 50 nM lead and demonstrate use of PbrR as a cell-free biosensor. We envision that our workflow could be applied to engineer any aTF.

  View details for DOI 10.1021/acssynbio.4c00456

  View details for PubMedID 39255329

• A frugal CRISPR kit for equitable and accessible education in gene editing and synthetic biology. Nature communications Collins, M., Lau, M. B., Ma, W., Shen, A., Wang, B., Cai, S., La Russa, M., Jewett, M. C., Qi, L. S. 2024; 15 (1): 6563

  Abstract

  Equitable and accessible education in life sciences, bioengineering, and synthetic biology is crucial for training the next generation of scientists, fostering transparency in public decision-making, and ensuring biotechnology can benefit a wide-ranging population. As a groundbreaking technology for genome engineering, CRISPR has transformed research and therapeutics. However, hands-on exposure to this technology in educational settings remains limited due to the extensive resources required for CRISPR experiments. Here, we develop CRISPRkit, an affordable kit designed for gene editing and regulation in high school education. CRISPRkit eliminates the need for specialized equipment, prioritizes biosafety, and utilizes cost-effective reagents. By integrating CRISPRi gene regulation, colorful chromoproteins, cell-free transcription-translation systems, smartphone-based quantification, and an in-house automated algorithm (CRISPectra), our kit offers an inexpensive (~$2) and user-friendly approach to performing and analyzing CRISPR experiments, without the need for a traditional laboratory setup. Experiments conducted by high school students in classroom settings highlight the kit's utility for reliable CRISPRkit experiments. Furthermore, CRISPRkit provides a modular and expandable platform for genome engineering, and we demonstrate its applications for controlling fluorescent proteins and metabolic pathways such as melanin production. We envision CRISPRkit will facilitate biotechnology education for communities of diverse socioeconomic and geographic backgrounds.

  View details for DOI 10.1038/s41467-024-50767-2

  View details for PubMedID 39095367

• At-Home, Cell-Free Synthetic Biology Education Modules for Transcriptional Regulation and Environmental Water Quality Monitoring. ACS synthetic biology Jung, J. K., Rasor, B. J., Rybnicky, G. A., Silverman, A. D., Standeven, J., Kuhn, R., Granito, T., Ekas, H. M., Wang, B. M., Karim, A. S., Lucks, J. B., Jewett, M. C. 2023

  Abstract

  As the field of synthetic biology expands, the need to grow and train science, technology, engineering, and math (STEM) practitioners is essential. However, the lack of access to hands-on demonstrations has led to inequalities of opportunity and practice. In addition, there is a gap in providing content that enables students to make their own bioengineered systems. To address these challenges, we develop four shelf-stable cell-free biosensing educational modules that work by simply adding water and DNA to freeze-dried crude extracts of non-pathogenic Escherichia coli. We introduce activities and supporting curricula to teach the structure and function of the lac operon, dose-responsive behavior, considerations for biosensor outputs, and a "build-your-own" activity for monitoring environmental contaminants in water. We piloted these modules with K-12 teachers and 130 high-school students in their classrooms─and at home─without professional laboratory equipment. This work promises to catalyze access to interactive synthetic biology education opportunities.

  View details for DOI 10.1021/acssynbio.3c00223

  View details for PubMedID 37699423

• At-home, cell-free synthetic biology education modules for transcriptional regulation and environmental water quality monitoring. bioRxiv : the preprint server for biology Jung, K. J., Rasor, B. J., Rybnicky, G. A., Silverman, A. D., Standeven, J., Kuhn, R., Granito, T., Ekas, H. M., Wang, B. M., Karim, A. S., Lucks, J. B., Jewett, M. C. 2023

  Abstract

  As the field of synthetic biology expands, the need to grow and train science, technology, engineering, and math (STEM) practitioners is essential. However, the lack of access to hands-on demonstrations has led to inequalities of opportunity and practice. In addition, there is a gap in providing content that enables students to make their own bioengineered systems. To address these challenges, we develop four shelf-stable cell-free biosensing educational modules that work by just-adding-water and DNA to freeze-dried crude extracts of Escherichia coli . We introduce activities and supporting curricula to teach the structure and function of the lac operon, dose-responsive behavior, considerations for biosensor outputs, and a 'build-your-own' activity for monitoring environmental contaminants in water. We piloted these modules with K-12 teachers and 130 high school students in their classrooms - and at home - without professional laboratory equipment or researcher oversight. This work promises to catalyze access to interactive synthetic biology education opportunities.

  View details for DOI 10.1101/2023.01.09.523248

  View details for PubMedID 36711593

  View details for PubMedCentralID PMC9881948