Josh is a postdoctoral fellow in Moh El-Naggar’s lab at the University of Southern California. His research focuses on using synthetic biology and protein engineering to control electron transport in biological systems. Josh is a member of the EBRC Student and Postdoc Association Board and works as a liaison to the education working group.
Ying-Chiang is a graduate student in the Donia Lab at Princeton University. He currently works on the capture and characterization of novel bioactive microbiome-derived peptides, particularly focusing on host-microbe interactions. The long term goal is to engineer therapeutic peptides from natural templates. Before coming to Princeton, Ying-Chiang completed his undergraduate work at Washington University in St. Louis where he worked in the Moon Lab and Virgin Lab. He then completed an MPH specializing in Global Health followed by an MEng.
Leah is a Biomedical Engineering Ph.D. student in the Daringer Lab at Rowan University. Her research focuses on engineering mammalian cell-based biosensors for the detection of extracellular ligands. Before starting graduate school, she received her undergraduate degree in Energy Engineering at The Pennsylvania State University, where she completed four internships and worked full-time for a year post graduation. If she’s not in the lab you can find her with her pug, Nugget.
Yan is a postdoctoral fellow at the California Institute of Technology. Her research focuses on engineering synthetic cells as the next-generation chassis to implement synthetic biology operations. Outside of research and EBRC, Yan is involved in the iGEM and science outreach efforts at Caltech to promote underrepresented groups’ engagement in synthetic biology.
As a graduate student in the Dunlop Lab at Boston University, Eric is developing optogenetic tools and feedback control strategies to improve how heterologous metabolic pathways integrate with native host cell physiologies. He is broadly interested in how synthetic biology and computer-aided techniques are being combined to accelerate the design of engineered microbes.
Friday, January 28th, 2021
12:00pm – 6:00pm Eastern | 9:00am – 3:00pm Pacific
***NOW VIRTUAL***
Note: due to the current COVID situation, this workshop has been changed to a virtual event.
The workshop will focus on drafting our technical research roadmap to advance engineering biology tools and technologies that respond to the urgency of the climate crisis and promote long-term sustainability.
This workshop is by invitation only.
Register Here
Registration closes January 18, 2022
Agenda: Coming Soon!
Workshop Objectives
For more information about the roadmap or workshop, please contact Sifang Chen (roadmap@ebrc.org)
Wednesday, October 27, 2021: 8:00 AM- 2:00 PM PDT
Thursday, October 28, 2021: 8:00 AM – 2:00 PM PDT
The annual EBRC Council Retreat and planning meeting will be held virtually October 27-28, 2021. This meeting is by invitation only and will be a mix of plenary discussions and extended breakout sessions.
Please register for the meeting and select your breakout session preferences, and indicate your interest in giving a Synergies lightning talk. We will organize breakout sessions based on your preferences, as well as obtaining a mix of academic and industry members, and WG representation. Throughout the meeting, we will identify and integrate opportunities to advance diversity, equity, and inclusion.
The EBRC Student and Postdoc Association is hosting a panel discussion called Follow the Money for researchers to understand the science funding pipeline for engineering and synthetic biology. Join us virtually on October 18th, 2021 (12:00-1:30pm ET | 9:00-10:30am PT) on Zoom by registering here. Our diverse panel includes a broad range of perspectives from the congressional, project management, and advocacy levels to provide a full-circle overview on how research priorities go from expert opinion to budget items to grants to research labs and back.
Click here to view our flyer.
Please contact Julie Ming Liang (julieliang2022@u.northwestern.edu) if you have any questions.
Climate and Sustainability Roadmap Virtual Writing Workshop 7
Monday, December 13th, 2021
12 pm – 3 pm ET | 9 am – 12 pm PT
Registration closes December 11th, 2021
Workshop Activities and Details Coming Soon!
We are excited to engage with EBRC members and the broader research community on this important roadmapping effort. We welcome workshop participation from academic, industry, and government scientists and engineers, research & development leadership, students and postdocs, and other stakeholders.
For more information about the roadmap or workshop, please contact Sifang Chen (schen@ebrc.org)
Climate and Sustainability Roadmap Virtual Writing Workshop 6
Wednesday, December 1st, 2021
11 am – 2 pm ET | 8 am – 11 am PT
Register Here
Registration closes November 29th, 2021
Workshop Activities and Details Coming Soon!
We are excited to engage with EBRC members and the broader research community on this important roadmapping effort. We welcome workshop participation from academic, industry, and government scientists and engineers, research & development leadership, students and postdocs, and other stakeholders.
For more information about the roadmap or workshop, please contact Sifang Chen (schen@ebrc.org)
Monday, November 15th, 2021
11:00am-2:00pm ET | 8:00am-11:00am PT
Register Here
Registration closes November 13th, 2021
Workshop Activities and Details Coming Soon!
We are excited to engage with EBRC members and the broader research community on this important roadmapping effort. We welcome workshop participation from academic, industry, and government scientists and engineers, research & development leadership, students and postdocs, and other stakeholders.
For more information about the roadmap or workshop, please contact Sifang Chen (schen@ebrc.org)
Thursday, September 30th, 2021
12:00pm-3:00pm ET | 9:00am-12:00pm PT
Register Here
Registration closes September 28th, 2021
Agenda: Click here
We are excited to engage with EBRC members and the broader research community on this important roadmapping effort. We welcome workshop participation from academic, industry, and government scientists and engineers, research & development leadership, students and postdocs, and other stakeholders.
This workshop will continue to refine the Roadmap’s technical themes and goals developed in previous workshops on engineering biology approaches to address climate, environmental, and sustainability challenges. In addition, participants will draft key technical objectives, milestones, bottlenecks, and potential solutions toward accomplishing those goals.
For more information about the roadmap or workshop, please contact Sifang Chen (schen@ebrc.org)
This virtual seminar series, held every Thursday (11-12 am US ET | 8-9 am US PT), will feature presentations by early-career professionals. You can view all seminars on Zoom or on the Moon Lab YouTube channel live stream. The seminar will be held almost every Thursday starting Jan 12, 2023. The schedule of speakers can be viewed here.
Zoom Webinar: https://us02web.zoom.us/j/88380504839
YouTube LIVE: https://www.youtube.com/channel/UCTkyl4Uw1NlpuLCeP4Zi8aA
Speaker Nominations: If you or someone you know is interested in speaking, click here for more information. Nominations will be accepted on a rolling basis.
Please contact Tae Seok Moon (tsmoon7@gmail.com) with any questions.
Thursday, September 9th, 2021
1:00pm-4:00pm ET | 10:00am-1:00pm PT
Registration closes September 7th, 2021
Agenda: Click here
We are excited to engage with EBRC members and the broader research community on this important roadmapping effort. We welcome workshop participation from academic, industry, and government scientists and engineers, research & development leadership, students and postdocs, and other stakeholders.
Building off the first two workshops focusing on climate and environmental challenges, this third workshop will broaden our scope to include discussions on concrete and bold engineering biology solutions for long-term sustainable development and use of natural resources, while iterating on the opportunities and goals put forth in the previous workshops.
For more information about the roadmap or workshop, please contact Sifang Chen (schen@ebrc.org)
Friday, August 20, 2021
11:00am-2:00pm ET | 8:00am-11:00am PT
Registration closes August 18, 2021
Agenda: Click here
We are excited to engage with EBRC members and the broader research community on this important roadmapping effort. We welcome workshop participation from academic, industry, and government scientists and engineers, research & development leadership, students and postdocs, and other stakeholders.
The workshop will provide an opportunity for participants to brainstorm, discuss, and put forth ambitious goals, breakthroughs, and milestones for advancing engineering biology to address climate and environmental challenges. This will be the second writing workshop and activities are anticipated to include reviewing and revising previously identified goals, along with additional writing.
For more information about the roadmap or workshop, please contact Sifang Chen (schen@ebrc.org)
Wednesday, July 28, 2021
2:00pm-5:00pm ET | 11:00am-2:00pm PT
Registration is now closed. For more information, please contact helix@ebrc.org
Agenda: Click here
We are excited to engage with EBRC members and the broader research community on this important roadmapping effort. We welcome workshop participation from academic, industry, and government scientists and engineers, research & development leadership, students and postdocs, and other stakeholders.
During the workshop, participants will:
For more information about the roadmap or workshop, please contact Sifang Chen (schen@ebrc.org)
Registration and abstract submission is now open for the upcoming HYBRID Central US Synthetic Biology Workshop, Sept 9-10th. If possible, the in-person portion of the workshop will be held at Northwestern University. Featured speakers include Caroline Ajo-Franklin from Rice and Huimin Zhao from U of Illinois Urbana-Champaign. The workshop website contains the most up-to-date information: http://centralsynbio.org/ We look forward to seeing you at this year’s Central US Synthetic Biology Workshop.
Registration and abstract submission for short talks and posters are open now! Register here!
Please register, regardless of if you will submit an abstract.
Please join us for an exciting seminar on May 21, 2021, from 4-5:00 PM ET. This is the final seminar in the 2021 EBRC Seminar Series.
Speaker abstracts are below. The seminar is open to all, so please feel free to share this information with your colleagues.
The seminar will be held on Zoom using the following link for all sessions:
Zoom link: https://berkeley.zoom.us/j/97626552307?pwd=alVlS3dXM0lZYklYeE9zVXljWUI0UT09
Meeting ID: 976 2655 2307
Passcode: EBRC2021
“Sequence-function analysis helps identify multiple pathways to enhance Phenylalanine Ammonia-Lyase (PAL) activity”
Nikhil Unni Nair (Tufts University)
Phenylalanine ammonia-lyases (PALs) non-oxidatively deaminate L-phenylalanine to trans-cinnamic acid (tCA) and are widely found associated with secondary metabolism in plants, bacteria, and fungi. Biocatalytic applications for natural product and fine chemical synthesis has driven the discovery, expression, characterization, and engineering of PALs. More recently, development of PALs for phenylketonuria (PKU) management and cancer therapy has further increased interest in engineering this class of enzymes. While there is a general understanding of how residues in the substrate-binding pocket contribute to specificity and turnover, led by rational mutagenesis studies, there is generally a poor understanding of how distal residues affect function. In general, outcomes from directed evolution can identify distal hotspots but there have only been two such studies with this enzyme. The first study resulted in modest improvement in activity whereas the other, conducted by us, identified residues within the active site only. Deep mutational scanning (DMS) can identify functional hotspots, and when coupled with directed
evolution can accelerate engineering campaigns, although there are few examples of this approach. Further, DMS can provide a comprehensive map of sequence–function relationships to explore the protein fitness landscapes, uncover functionally relevant sites, improve molecular energy functions, and identify beneficial combinations of mutations for protein engineering. Though extensive body of research exists on function, structure, mechanism of PAL, a systematic study exploring the sequence-function space has not been attempted.
Previously, we developed a growth-coupled enrichment for rapid screening of high-activity variants of AvPAL* (used to formulate the PKU drug Pegvaliase®) in E. coli. After a single round of directed evolution using this growth-coupled enrichment, we identified 2 active site mutations improved kcat < 2-fold. However, the sequence-function fitness landscape of AvPAL* remains to be explored. In this study, we achieve several outcomes. First, we obtained the detailed sequence-function landscape of PAL, to date, using DMS, identifying >60 mutational hotspots. Next, we picked seven sites for comprehensive single and multi-site saturation mutagenesis and we identified multi-site mutations with ~2.5-fold improvement in the kcat (and >3-fold increase in catalytic efficiency). We then explored the epistatic effect of these mutations, uncovering positive, neutral, and negative interactions among distal and proximal sites. Finally, to understand the mechanistic role of key mutations in hyperactive variants, we performed modelling studies and concluded that there are multiple pathways to enhance PAL catalytic activity, including, decreased root mean square fluctuation (RMSF) of substrate in the active site, greater proximity of the substrate to catalytic residues, and facilitated diffusion of the substrate to the active site, among others. In summary, this study significantly advances basic and applied enzymology of PALs, a heretofore understudied class of enzymes with a wide array of applications.
Development of a yeast-based assay for bioavailable phosphorous
Heather Shepherd (University of Notre Dame)
Preventing eutrophication of inland freshwater ecosystems requires quantifying the phosphorous (P) content of the streams and rivers that feed them. Typical methods for measuring P assess soluble reactive P (SRP) or total P (TP) and require expensive analytical techniques that produce hazardous waste. Here we present a novel method for measuring the more relevant bioavailable P (BAP); this assay utilizes the growth of familiar baker’s yeast, avoids production of hazardous waste, and reduces cost relative to measurements of SRP and TP. The yeast BAP (yBAP) assay takes advantage of the observation that yeast density at saturating growth increases linearly with provided P. We show that this relationship can be used to measure P in freshwater in concentration ranges relevant to eutrophication. In addition, we measured yBAP in water containing known amount of fertilizer and in samples from agricultural waterways. We observed that the majority of yBAP values were between those obtained from standard SRP and TP measurements, demonstrating that the assay is compatible with real-world settings. The cost-effective and nonhazardous nature of the yeast-based assay suggests that it could have utility in a range of settings, offering added insight to identify water systems at risk of eutrophication from excess phosphorus.
Engineering alternative degradation tags for synthetic circuits
Prajakta Jadhav (South Dakota State University)
The goal in synthetic biology is to build robust synthetic circuits in bacteria that are dynamic, highly regulated, and result in a unified response. In the last 20 years, the synthetic biology field has effectively leveraged transcriptional (RNA production) and translational (protein production) controls. However, protein degradation plays an important role in determining the half-life of proteins and regulating biological systems. Amino acid degradation tags are exploited to avoid a reliance on cell division for protein dilution and to build dynamic circuits. The most leveraged
degradation tag in E. coli is the ssrA-tag, which is an 11-amino acid sequence that primarily target proteins for degradation by the ClpXP proteolytic system. However, the use of the ssrA-tag limits scalability and complexity especially in synthetic oscillators in bacteria due to itsmultiple proteolytic target recognition signals. Our goal is to build orthogonal oscillators that utilize degradation tags targeting to multiple proteases with no crosstalk. In this study, we aremodifying the ssrA-tag to change the substrate affinity and degradation rate and produce new
synthetic oscillators. We hypothesize that changing the degradation rate can alter the output signal of the oscillator. We have tailored the ssrA-tag to reduce crosstalk between proteolytic systems to increase robustness and developed a variety of degradation tags. While many of these tags exhibited decreased or no change in degradation rates, two depicted significant increase. We further tested interesting candidates for crosstalk between proteolytic systems and identified a tag that display minimum to no crosstalk with other proteolytic systems. We aim to build and compare the output signals of different oscillators with novel tags in batch cultures and at the single-cell level. The design and implementation of these novel degradation tags will enable development of biological building blocks for increased complexity in synthetic circuits.
Please join us for an exciting seminar on May 13, 2021, from 2-3:30 PM ET. This is the second of three seminars in the 2021 EBRC Seminar Series.
Speaker abstracts are below. The seminar is open to all, so please feel free to share this information with your colleagues.
The seminar will be held on Zoom using the following link for all sessions:
Zoom link: https://berkeley.zoom.us/j/97626552307?pwd=alVlS3dXM0lZYklYeE9zVXljWUI0UT09
Meeting ID: 976 2655 2307
Passcode: EBRC2021
TBD
Aindrila Mukhopadhyay (Lawrence Berkeley National Lab)
TBD
“Non-canonical crRNAs derived from host transcripts enable multiplexable RNA detection by Cas9”
Chunlei Jiao (Helmholtz Institute for RNA-based Infection Research)
CRISPR nucleases are guided by CRISPR RNAs (crRNAs) that are naturally derived from CRISPR arrays. Here, we discovered that crRNAs could derive from cellular transcripts outside of the CRISPR-Cas locus in Campylobacter jejuni, and we exploit this discovery to achieve a novel means of multiplexed RNA detection. The discovery came from sequencing RNAs that preferentially bound to the Cas9 in C. jejuni (CjeCas9), revealing an unexpected set of RNAs that shared a motif complementary to the anti-repeat of the system’s tracrRNA. The size and composition of these bound RNAs resembled that of crRNAs, indicating that the full-length version of these RNAs base pair with the anti-repeat portion of the tracrRNA and undergo processing to form crRNA-like RNAs. We call these processed RNAs non-canonical crRNAs (ncrRNAs). Using a cell-free transcription-translation (TXTL) system, we found that some ncrRNAs could drive efficient and sequence-specific DNA cleavage by CjeCas9 and its natural tracrRNA. Given the known sequence flexibility within the repeat:anti-repeat stem for single- guide RNAs, we hypothesized that the anti-repeat portion of the tracRNA could be reprogrammed to convert any RNA-of-interest into a functional ncrRNA that guides Cas9 to its DNA target. Using DNA cleavage assays in TXTL and in E. coli, we found that reprogrammed
tracrRNAs(Rptrs) designed to pair with different regions of an mRNA yielded efficient DNA cleavage not only for the CjeCas9 but also for the S. pyogenes Cas9 and the Streptococcus thermophilus CRISPR1 Cas9. Finally, based on the capability of Rptrs to link any RNA-of-interest to sequence-specific DNA cleavage, we established a multiplexed RNA diagnostic platform called LEOPARD (Leveraging Engineered tracrRNAs and On-target DNAs for PArallel RNA Detection). With LEOPARD, we achieved multiplexed detection of RNAs from different viruses including SARS-CoV-2 and other respiratory viruses in one reaction. We further distinguished SARS-CoV-2 and its D614G variant with single-nucleotide specificity in patient samples. These findings establish a previously unknown source of crRNAs and demonstrate the practical utility of LEOPARD for detecting numerous biomarkers in one test.
“Developing a mathematical framework for controlling complex biological systems”
Marcella Gomez (UC Santa Cruz)
In this talk, we refer to the achievement of an intended and predicted response in a biological system as controlling biology. Such efforts are often guided by classical mechanistic models from first principles. The level of complexity of these systems makes it extremely difficult to develop mechanistic models that can account for all possible interactions and predict biological response. To overcome these challenges, we propose to move away from first principal methods and instead identify key leverage points in the targeted biological pathways that can be directly up or down regulated by external signaling molecules. These molecules can be controlled by a feedback algorithm and delivered in situ by a bioelectronic device. In recent work, we successfully implemented feedback control on human‐induced pluripotent stem cells (hiPSCs) to regulate the cell’s resting potential known to affect cell physiology and functions such as proliferation, differentiation, migration, and apoptosis, as well as cell–cell communication and large‐scale morphogenesis. This was achieved without use of a model nor training data. We further outline an approach to extend the method to more complex biological systems. In particular, we consider the task of accelerating wound healing.
“The Promoter Calculator – A Sequence-to-Function Biophysical Model of Transcriptional Initiation for Sigma70 Promoters with Any Sequence”
Travis La Fleur (Penn State)
Engineering synthetic promoters with precision control has remained a challenge due to our inability to predict how a promoter’s sequence and DNA context controls its function and mRNA output. Here, we developed an accurate sequence-to-function model of transcriptional initiation that enables the automated design of synthetic promoters and the a priori prediction of cryptic promoters within natural systems – both of which are needed to advance Synthetic Biology towards genome-scale functional design. To do this, we combined oligopool synthesis, library-
based cloning, and next-generation sequencing to construct and characterize 14,206 rationally designed sigma70 promoters in vitro. Measurements include transcriptional start site frequencies and overall mRNA levels. This approach enables highly-parallel characterization of constitutive promoter activity without confounding factors such as unintentional transcriptional regulation, non-sigma70 activity, and mRNA decay. These measurements, in combination with machine learning, were used to parameterize a thermodynamic model quantifying the
interactions controlling transcription rate. For demonstration, the “Promoter Calculator” was used to accomplish three major tasks – accurate prediction of thousands of sigma70 promoters across various conditions, the de novo design of novel sigma70 promoters, and the identification of cryptic promoters internal to a genetic circuit. 4,350 highly non-repetitive promoters and 6,165 genome-integrated promoters characterized in vivo were accurately predicted by the model with Spearman Rank-Order Coefficients of .68 and .78, respectively. Promoters designed de novo using the Promoter Calculator were characterized in vivo exhibiting a 683-fold range in expression resulting in a Pearson Correlation Coefficient of .85. The Promoter Calculator was used to analyze a genetic circuit containing 11 circuit promoters and 29 cryptic promoters. The model identified 29 sigma70 promoters out of 40 observed promoters (72.5% accurate), including 10/11 circuit promoters and 19/29 cryptic promoters. The Promoter Calculator facilitates context-aware, rational promoter design without relying on a fixed table of pre-characterized sequences while serving as a powerful tool in promoter identification.