Mock Orange - Poems by Anne Osbourn

Please join us in congratulating OpenPlant PI Prof Anne Osbourn on the publication of her book of poetry entitled Mock Orange:

MOCK ORANGE by ANNE OSBOURN  | SPM Publications, Poetry

Third Prize Winner, Sentinel Poetry Book Competition 2018

“Mock Orange is a collection of poems in which Anne Osbourn attempts to order her life and her origins and to try to understand how and why she became a scientist, specifically a plant biologist. From early childhood she has tried to make sense of the world through plants. In mid-eighteenth century Sweden Linnaeus, the father of modern taxonomy, spent his life trying to understand his Maker through the classification of plants. Osbourn’s poetry encompasses Linnaeus’s adventures and experiences and his fascination with living things. Mock Orange is therefore about journeys from origins, both personal and global, in which negotiations between scientific and non-scientific languages and points of view form a central theme.”

Post Doctoral Research Assistant, University of Edinburgh

Post Doctoral Research Assistant  – Experiments of Topologically Active Polymers & DNA Origami
From June 2021 or soon after to June 2024 (3 years) & potentially extendible
Salary high-end UE07 ~ £41k/year

We are looking for an experienced and motivated PDRA with background in experimental soft matter, polymer physics and/or molecular biology. The ideal candidate should have direct expertise on DNA biophysics and/or rheology of complex fluids. 

The project involves developing and performing experiments on polymers that can change topology in time and that are based on DNA. It also involves experiments with DNA origami and super-resolution microscopy. The aim of the project is to understand the rheological properties of polymeric materials and complex fluids made of “topologically active” polymers and DNA origami. 

This position is for a senior PDRA, who is expected to be semi-independent, have supervision responsibilities on a day-to-day basis and to propose and lead side-projects.

More info: https://www.vacancies.ed.ac.uk/pls/corehrrecruit/erq_jobspec_version_4.jobspec?p_id=052998

Contact: Davide Michieletto, davide.michieletto@ed.ac.uk
Group website: www2.ph.ed.ac.uk/~dmichiel/eutopia.unitn.eu

CtoD: From Cells to Droplets

The aim and rationale

Real biological systems are complicated by the massive number of components and highly complex cellular network. Establishing a cell-like environment in vitro would allow us to disentangle basic mechanisms underlying cellular networks under well-controlled settings. We are a small but interdisciplinary team between the Department of Chemistry and the Department of Plant Sciences, aiming to apply microfluidic droplet system to studying physiological processes in plants, transforming the objects of research from plant tissues into artificial cells.

Microdroplets generation and in-droplet protein expression for mimicking physiological pathways.

The project and more

In this project, we aimed to develop an artificial cellular system using microfluidic droplets that allows the in vitro expression of proteins from plants in a compartmentalised cell-like environment. The biosynthetic pathway of a class of plant pigments, betalains, was used as a simple model to test the methodology, and the TNT® SP6 High-Yield Wheat Germ Protein Expression System from Promega was selected as a reliable in vitro protein expression system. The cell-free protein expression components together with betalain biosynthetic genes were enclosed in microdroplets and intermediate and final products measured.

Betalain biosynthetic pathway from L-Tyrosine to two groups of betalains. DOPA, 3,4 dihydroxyphenylalanine. DODA, 4,5 DOPA dioxygenase. cDOPA5GT, cyclo-DOPA 5-O glucosyltransferase.

Up to this point, we have tested the expression of various proteins in microdroplets. Betalain biosynthetic enzymes were produced and confirmed by fluorescent protein fusions in microdroplets. However, simply co-expressing enzymes from the betalain biosynthetic pathway in the cell-free protein expression solution was not sufficient to drive detectable production of pigment molecules. Following these, optimisation of buffer composition was made and various substrates fed into the system, which led to realisation of parts of the pathway in the in vitro environment.

In addition to the project itself, the various events and training organised by OpenPlant/Biomaker were even more remarkable. The annual OpenPlant Forum gathered an incredibly diverse range of subjects and the low-cost DIY instruments for automating laboratory experiments were mind-blowing.

Future perspective

The OpenPlant/Biomaker project has built solid foundation and connection for us to move forward in the future. Communication between cells plays a central role in many metabolic pathways of multicellular organisms, yet it remains difficult to reproduce in vitro. To this end, an artificial multicellular system with droplet-to-droplet communication will be explored to mimic metabolic pathways which rely on the communication between cells. This platform would open more opportunities for synthetic biology and allow hypotheses and models based on cell-to-cell communications to be tested in vitro.

Last but not least, we thank the OpenPlant/Biomaker teams for the wonderful events and generous help that they have provided!

Written by Zhengao Di, University of Cambridge

Developing an open and affordable 3D bioprinter

Background

Friends had Central Perk cafe, the gang from How I Met Your Mother had MacLaren’s Pub and we have Charlie’s Pizza joint. What started as a trivial discussion about printing human organs over a stone-baked Margherita quickly evolved into us applying for a bid to participate in the BioMaker Challenge, with the objective of Developing an open & affordable 3D bioprinter. 

The beginnings of 3D printing date back to the 1980’s, when it was commonly known as its more mouthful synonym stereolithography (SLA), a technology pioneered by Charles W. Hull. Revolutionized with SLA, one could translate a 3D design from a data file into a physical object, in a relatively short time. The household notion of 3D printing is testament to the success of SLA, which originated from the company 3D Systems Corporation. 

Fast forward 40 years, my neighbours are at home 3D printing little elves for their garden, my cousin is buying a 2-story building printed by an enormous SLA machine, my doctor is offering his patients custom 3D printed ear implants, heart valves or bones - SLA has already infiltrated many aspects of our lives, economy and medical care. Much of this progress came about because of the RepRap project, whose aim was to create an open-source 3D printer capable of printing most of the parts needed to replicate itself, making it cheap enough for hobbyists. Today’s consumer 3D printer companies, and the widespread use of 3D printing, grew out of the RepRap hobbyist community. 

The field of regenerative medicine could hugely benefit from 3D printing techniques to offer personalized medicine to its patients, by for example printing organs made of one’s own cells. However, these applications are challenged by the complex architecture of human organs, the difficulties of supporting living cells, introducing foreign materials in a human body and working at a microscale. 

Addressing these issues, companies and academic researchers have built 3D printing platforms, specialized for biological materials, hence 3D bioprinting. Not surprisingly, these come at a prohibitive cost, up to £200K for the printer only.

BioMaker Challenge

During our weekly pizza gatherings we talked about how it would be great to have an open-source bioprinting project, along the same lines as the original RepRap project. It could make the technology more accessible and bring printing human organs for transplantation a step closer. None of our backgrounds were in bioprinting, but our combined skillsets did range from engineering and programming to cell biology, with diverse backgrounds in start-ups, industry and academia. The Biomaker challenge would be a great way to work together on a project where we could contribute something to the 3D printing community, and learn a lot along the way. Plus it would be fun! 

Bertie the 3D bioprinter.

So as part of the BioMaker Challenge, we set out to address the lack of reasonably priced open-source 3D bioprinters in the market by Developing an open & affordable 3D bioprinter. With this project still in its infancy, working with human cells was out of the scope due to ethical and practical reasons. Nevertheless, we attempted to design our 3D printer to potentially print corals, an application with a shorter timeline, more amenable to working in the absence of a biological laboratory and with direct benefits to a problem close to our hearts, the depreciation of the ocean’s corals. Our project branched into two components outlined below: 

1. Converting an existing open-source 3D printer into a 3D bioprinter capable of extruding biomaterial.

Our inspiration to address this first problem is drawn from a paper by an American group (Push et., 2018), in which they describe the design of a syringe pump large volume extruder (LVE) of low cost, compatible with printing biomaterial. This involves the purchase of standard materials (e.g: 60mL syringe, bolt nuts) and the 3D printing of other assembly components with a PLA extruder from a standard 3D printer. The newly assembled LVE would then replace the printer’s standard PLA extruder and allow the printing of biomaterials. 

With this in mind, many hours of research and discussions went into selecting the right 3D printer as a foundation for our bioprinter - we were certain about two things: it had to be open-sourced and stay within a reasonable budget. We finally settled on the RepRap Ender 5 3D-printer. In addition to the syringe extruder, we were interested in trying to integrate a peristaltic pump in our system because it would allow us to extrude multiple bio-inks at once. 

Adding a valving architecture between the reservoirs and the pump enables the switching of the reservoir that bioinks are aspirated from on the go, thereby enabling the interlinking of multiple inks of different cell types in 3D space. 

After a few months of putting all the hardware together and with a sprinkle of software, Bertie the Bioprinter was born!

2. Developing a bioink suitable for sustaining living cells.

Regardless of the organism in question, a bioink should: 

a) Contain the right nutritional environment for specialized cells.

b) Maintain a single-cell suspension (prevent cells from clumping together and/or from dying from being separated from their friends).

c) Provide a physical scaffold once printed to keep the shape of the 3D model.

d) Use materials that are safe to insert in the receiving environment. 

Printing human cells was out of the scope of Biomaker, so we initially turned our attention to another challenging bioprinting application, trying to 3D print coral (which not all of us knew was a living organism!). Coral numbers and health have declined rapidly as a result of environmental conditions and their slow growth cycle. If coral could be 3D bioprinted, perhaps it could be transplanted back into reefs to bring them back to life.

We dove into the scientific literature and identified two such bioink formulas, which are, in theory, capable of supporting the life cycle of corals and providing an “ocean-friendly” physical scaffold. It turned out however that coral species are high maintenance, requiring expensive aquariums and daily care. Even the most experienced coral growers find it challenging to maintain a community of the families of coral that were the right size to fit our bioprinter. We realised that we didn’t have the funds or the time to manage coral during the Biomaker Project, but we did make connections with coral researchers who we hope to work with on this project in the longer term. 

Printed phytoplankton using Bertha the bioprinter.

We were however determined to evaluate the efficacy of our bioinks, and didn’t let coral’s needy demands deter us. One of the bioinks we identified is based on the chemical interaction of sodium alginate and calcium chloride, which create a solidified gel upon contact. At the concentration used, these chemicals didn’t affect the survival of phytoplankton, a single cell organism we chose to work with for its relative low cost, ease of maintenance and ability to survive as single cells. 

By the end of the project we were able to print a very slimy collection of phytoplankton, and affectionately named it Bertha. Bertha still sits in someone’s fridge for posterity.

Future

The BioMaker Challenge was an incredible opportunity for us to concretise an idea and start along the path to our long term goal. We also got to meet a wonderful community of makers and with their help and feedback we hope to further refine Bertie the Bioprinter. Moving forward, we plan to improve Bertie’s performance in terms of printing precision and its ability to run complex modes (e.g: multiple bioinks printing at once), as well as make him suitable for sterile work (e.g: ventilation, UV lights). We also plan to develop  application specific bioinks for Bertie to print. 

We’d like to take this opportunity to thank the BioMaker Organisers (special thanks to Alexandra, Jenny & Jim) for arranging this event, Professor Ludovic Vallier (Cambridge Stem Cell Institute, Jeffrey Cheah Biomedical Centre) for supporting our ideas and administrative needs as well as the BioMakeSpace Cambridge who welcomed us in their community.  Thanks to the BioMaker Challenge, we are now one small step closer to 3D printing organs, and in the meantime, Bertie is on standby for printing COVID-19 related materials (with the original PLA extruder) if need be. 

Julie, Sebastian, Monica, Robin, Ben & Tejas

PS, For more details of the project, check out our Hackster :-)

OpenCM - an open framework for single cell manipulation

Introduction

Microscopic studies of single cells, their dynamics, signalling and behaviour require a whole toolbox of different microscopic techniques, highly specific gadgets and methods that can easily exceed the budget for new lab equipment.

OpenCM team (left to right): Maziyar Jalaal, Nico Schramma, Stephanie Höhn and Kyriacos Leptos

The open technology community is continually developing new, innovative and inexpensive solutions for easy-to-build microscopes and other equipment, aiming to democratize microscopy and enhancing the applicability of custom-built devices to approach novel questions. The OpenCM team aimed to provide one of the solutions needed to make open-source microscopes even more utile, namely a setup for micromanipulation.

Multi-axis micromanipulation is used for in-vitro fertilization, imaging of highly motile cells, microinjections, patch-clamp methods, mechanical probing of cells with micropipettes and is - simply put - the microscopists tiny helping hand to interact with the sample mechanically.

The project

Key to the OpenCM project is the close collaboration with Benedict Diederich and René Lachmann from the UC2 group, which started after the OpenPlant forum in July 2019. UC2 builds 3D-printed modular optical elements, which can be easily assembled into various microscopes. The OpenCM project aims to make this system even more versatile, by adding a module for micromanipulation.

During the BioMaker challenge the team was able to design the first prototype two-axis micromanipulator, which fitted the UC2 system and was controlled via Arduino. The team followed the online documentation of their collaborators in order to 3D-print a microscope, which is controlled with an open source Python based GUI on a Raspberry Pi. The team was among the first to reproduce the UC2 microscope and helped evaluate and enhance their current quality of the online documentation.

In the end, the team was able to build a fully integrated microscope and micromanipulation system and to showcase it at the BioMaker Fayre. The most important outcome of the BioMaker Challenge is establishing a very close collaboration with the UC2 project, which is still persisting.

Future work

The team plans to undertake the final steps in developing a 3-axis micromanipulator (OpenCM2). The hardware will be integrated into the UC2 system, and free software will be provided for position controlling with a wireless joystick and the UC2 GUI via Raspberry PI and wifi controllers.  

First prototype as presented at the BioMaker Fayre 2019, an inverted microscope with 2 axis micro
manipulator.

The project will continue in close collaboration with the laboratory of UC2 in Jena, Germany, and the financial support in the context of the programme “Creativity and Studies” of the University of Gottingen by the AKB Stiftung, a non-profit foundation of the Büchting Family (secured by Nico Schramma and Björn Kscheschinski).

Furthermore, the team found the UC2 setup very useful to provide hands-on experience at the intersection between physics, mechanical engineering, microscopy and cell biology. Hence, as a side project, the team also started preparing workshops for children and students.

In a joint effort, and with the financial help of the UK's Women Engineering Society and the Public Engagement office at the University of Cambridge, the members of UC2 and OpenCM participate at the Women in Engineering Society 100 Violets Challenge in order to give a first "Optics for Everybody" workshop. Unfortunately, due to the pandemic, the exhibition at the Brunel Museum in London has been postponed.

During the BioMaker challenge and driven by the community's open-mindedness, the team was encouraged to branch out ideas, communicate openly, and team up with others to find and combine new and innovative solutions. In the end, the BioMaker spirit inspired the OpenCM team to reach out to the broader community.

Acknowledgments

We thank David Page-Croft for fruitful discussions and his help with 3D printing. We also thank Benedict Diederich, René Lachmann and Barbora Marsikova from the UC2-Project, and Alexandra Ting, Dieuwertje van Esse - van der Does and Jim Haseloff from the BioMaker Challenge.

Written by Maziyar Jalaal, Nico Schramma, Stephanie Höhn and Kyriacos Leptos.

References

Diederich, Benedict, et al. "UC2-A Versatile and Customizable low-cost 3D-printed Optical Open-Standard for microscopic imaging." bioRxiv (2020).

Research Assistant/ Lab Manager position in Chemical Biotechnology, University of Edinburgh

3.5-year Research Assistant / Lab Manager position in Chemical Biotechnology (Wallace Lab - IQB3)

We are looking for a highly-motivated MSc/PhD-level Research Assistant to join our lab in IQB3 from Dec/Jan. The role will be primarily research based and will continue our work on the combined use of chemical and biological tools in microbes for the sustainable production of industrial small molecules. This will include the assembly and optimisation of biosynthetic pathways in bacteria, the screening of new biocompatible catalysts, and the analysis of novel metabolites using a variety of analytical techniques. The post will also include a significant supervisory role, including the management of day-to-day research activities in the group, organisation of lab strains, coordination of group meetings and seminars.

Applicants can contact Stephen directly (stephen.wallace@ed.ac.uk) including a CV and cover letter, or apply directly using the link below.

Closing Date: 12th October 2020

Link here: https://www.jobs.ac.uk/job/CBP647/research-assistant

Website: http://wallacelab.bio.ed.ac.uk 

Twitter: @Dr_StephenW and @Wallace_Lab

Research Associate in Protein Design and Synthetic Biology, University of Bristol

Research Associate in Protein Design and Synthetic Biology

With Dr Fabio Parmeggiani, University of Bristol  

The group of Dr Fabio Parmeggiani is seeking a talented and enthusiastic postdoc interested in protein design and synthetic biology. The project will be focused on designing custom modular protein architectures as robust and reliable scaffolds for the development of novel functional protein-based nanomaterials and multivalent ligands to modulate cell response.

The aim of the project is to develop a robust, fast and high throughput design platform to rapidly build new proteins and complexes. The researcher will be responsible for developing the experimental part of the pipeline (involving DNA assembly, protein expression and purification, validation of designs through biophysical and structural characterization) and spearhead applications in nanomaterials and cell biology. The position would be best suited to a researcher with a keen interest in applying de novo protein design in synthetic biology.  Essential skills for this role would include: design, construction and expression of synthetic genes in E. coli; biochemical and biophysical characterisation of proteins; strong structural biology expertise, in particular with cryo-electron microscopy and x-ray crystallography.

Closing date: 14 October 2020 

More information can be found here.

Plant Scientist position at Phytoform Labs

The Company

At Phytoform Labs Ltd we believe in sustainable agriculture and nutrition for all. We see a lack of innovation in the agricultural space around the crops themselves, so we are building a multidisciplinary team to help rectify the situation. We have exciting growth plans over the next year and are looking for a talented plant scientist to support our R&D pipeline and genome engineering projects. We are based at the world leading plant science institute Rothamsted Research just outside of London, United Kingdom.

The Role

If you are looking for a new challenge and like working with a multidisciplinary team then this job is for you. We are looking for a motivated core team member to work on Phytoform Labs R&D and genome editing efforts. You will quantify and analyse multiple aspects of crop genetics as well as breed new crops with footprint-free genome editing. If you have previously been part of an academic or industry team, but now would like to work in a fast-paced start-up environment and develop a project from the ground up then you would be a great fit.

More information

Design of low cost reactors for molecular diagnosis and bio engineering.

Jim Haseloff (Plant Sciences, University of Cambridge), Francesco Ciriello (Mathworks, Cambridge) and Maziyar Jalaal (Department of Applied Mathematics and Theoretical Physics, University of Cambridge).

OpenPlant Biomaker Design2020

Modern biology has exploited enzymes and clever reaction systems to produce an ever-growing range of diagnostics, environmental assays, DNA assembly and tools for education and research. Many of these molecular reactions are cheap and accessible, but require instrumentation for precise temperature regulation, programmed control over time and quantitative readout. Commercially available instruments are expensive, employing milled metal components for temperature control of hundreds of samples. There are many cases where low cost (<<£100) reactors would be very useful, even if sacrifices were made for the scale of sample numbers or flexibility or speed of temperature control.

We have taken on the challenge of exploring new open solutions for ultra-low cost micro-incubators. At the beginning of the COVID-19 lockdown, we set up an international Biomaker expert group to tackle this. Currently, around 75 participants and observers are connected via web exchange and regular teleconferencing. We have built: (i) a programmable rig for testing prototypes. The integration of no-code programming and programmable touchscreen means that the instrument can be highly flexible, and easily repurposed. (ii) This has inspired the assembly of a small control unit that can be used for low-cost implementation, after the testing process. (iii) Quantitative models for air flow and heat transfer to inform new designs and computer control. Air heating/transfer can potentially provide a cheaper and more accessible route to incubator design, compared to the conventional use of heaters attached to milled aluminium blocks. (iv) We have incorporated cheap heating hardware into custom 3D printed housings and are testing the designs.

Quantitative modelling

Figure 1: The control system for the simulated instrument (©MathWorks).

3D printing allows the design and low-cost testing of possible solutions that can include high levels of geometric complexity. One of the challenges with building 3D printed incubators is the prospect of softening or melting of thermoplastics - in our case, we'd ideally like to have a working range of zero to 95 degrees C - to encompass most commonly used DNA procedures. One of the expert group, Carlo Quinonez, has pioneered the use of 3D printing techniques to experiment with complex manifolds and flues for microscope-based cell culture incubators - and to ultimately build a high-temperature (190 degrees C) convection oven using heat resistant ULTEM 1010 plastic (https://studiofathom.com/blog/pyrahttps://www.instructables.com/id/Shape-of-things-to-come/), and we have taken inspiration from this approach.

LAMP, PCR and other incubator reaction protocols require precise and often dynamic temperature control. The thermal response of a reaction vessel, and as a result the performance of an incubator, depends on a variety of factors. These include:

•   the material properties of the device components, e.g. their thermal conductivity, density and specific heat capacity;

•   and of the fluid surrounding them, e.g. its thermal diffusivity, viscosity, density;

•   the geometry of these components, e.g. their surface areas, volumes, thicknesses, shapes, arrangements;

•   the ensuing flow patterns that result from the convection of heat along their surfaces, e.g. typified by spatially-varying flow velocities, temperature gradients and flow regimes (laminar vs turbulent flows, forced vs natural convection).

The complex nature of convective flow patterns presents many challenges to the successful design of an incubator. The correct estimation of heat transfer coefficients for components often requires detailed experimental or numerical studies. Fortunately, there is a wealth of literature when it comes to the documentation of these studies (Holman 2009) and results for simplified geometries can be drawn upon to obtain sensible estimates for these coefficients. These idealisations can inform design choices at an early stage. Relying on these, a modelling framework can be established that can successively be improved, moving forward, by a combination of hardware testing and supporting high-fidelity FEA and CFD simulations.

To build a theoretical foundation for the design, Francesco Ciriello drafted a simplified thermal model of an incubator in Simscape. Simscape is a modelling tool within the Mathworks Simulink product family that allows for the rapid design of 1D lumped-mass models for multi-domain systems.

Figure 2: Model variants were used for the controller and plants to systematically assess the impact of specific design choices on the thermal response of the samples (©MathWorks).

Lumped thermal masses are used to idealise the samples, the air in the chamber and the reaction chamber walls. Heat transfer coefficients and resistances are extracted from Holman’s book (Heat Transfer. 10th Edition, McGraw-Hill, New York, 2009) Heat Transfer book for different geometries and scenarios. The simulated instrument can now be used to experiment with design choices and controller design and tuning (Figure 1). Simulink model variants have been particularly helpful at enabling these comparisons (Figure 2).

The simulations are encouraging us to explore different parameter spaces and question traditional design paradigms, e.g. the use of resistance heaters strapped to milled aluminium blocks. Simulations are also empowering when access to hardware resources is limited. The combination of Model-Based Design, rapid prototyping with additive manufacturing technologies and low-cost electronics are a powerful enabler for collaboration on open-science projects and can democratise access to a wealth of laboratory resources for biological research and medical diagnostics.

Mazi Jalaal has used Computational Fluid Dynamics to explore important parameters, such as heat transfer coefficients, in the model explained above. He used the CDF package Comsol to simulate the fluid flow and heat transfer inside an incubator. Figure 3 shows an example of the simulations, where warm air entering the closed incubator heats a series of reaction vessels.

Figure 3: Heating eight reaction vessels inside an incubator. Colours show the temperature field

Mazi Jalaal has used Computational Fluid Dynamics to explore important parameters, such as heat transfer coefficients, in the model explained above. He used the CDF package Comsol to simulate the fluid flow and heat transfer inside an incubator. Figure 3 shows an example of the simulations, where warm air entering the closed incubator heats a series of reaction vessels.

Figure 4. Test-rig and compact control unit

Test-rig and compact control unit

One challenge was to build a test-rig that would combine the flexibility of microcontroller-based electronics with code-free programming in XOD - and speed up the development and testing of control routines and graphical user interfaces. Jim Haseloff has built a testing device with integrated programmable touchscreen to simplify design and prototyping of the instrumentation. The screen manufacturers provide software that allows drag-and-drop programming of multi-screen user interfaces, with a corresponding set of Arduino libraries. Further, XOD programmers have converted the Arduino IDE libraries to XOD libraries, with a set of nodes that allow simple visual coding of interactions between the microcontroller and screen. For example, the touchscreen can provide multiple sets of switches, knobs, gauges and level settings and sophisticated display of parameter settings or sensor measurements - all quickly reconfigurable in software. The test-rig provides a base for controlling and monitoring prototype instruments. In order to then adopt the interface for real-world instrumentation, we're experimenting with compact lower-cost hardware that can be used to build standalone instruments.

Figure 5. Components chosen for prototype, with 200µL 8-tube strip for reference.

Figure 6. 3D design for reactor vessel (left) and printed prototype with hardware installed (right).

Components for a simple prototype microreactor

Jim Haseloff selected a simple set of low-cost hardware components: PTC heater, 40mm fans, 40mm heatsinks, MOSFET modules for computer controlled switching, DS18B20 pre-calibrated digital temperature sensors and 12V DC power supply (total ~$25). These were assembled in 3D printed housings that had been designed in Autodesk Fusion 360, printed on an Ultimaker S3 printer, and were tested using the programmable test-rig assembled for this purpose.

The prototype incubator was designed for heating small (0.2 µL) reaction tubes to around 60o Celsius for LAMP diagnostics. To minimise costs and complexity, a positive temperature coefficient (PTC) heater was used as a heat source. PTC heaters provide resistive heating. When cool, the device draw a relatively large current. As the temperature rises, the resistance increases - resulting in a fall in current. This continues until the heater reaches a maximum temperature. Therefore PTC heaters do not overheat, and this simplifies the design of both the prototype control system and vessel. The prototype was intended to exploit air heating (rather than metal block heating) and vessel shape and baffles to maximise efficient heat exchange with the incubator.

Figure 7. Real-time monitoring through digital temperature sensors. Upper right panel shows the plotted output of DS18B20 temperature sensors in a prototype incubator, showing heating from room temperature (24 degrees C) to a target of 60 degrees. The heater output was controlled from XOD, and the temperature increase was logged onscreen. (The x-axis graticule marks are 20 secs apart).

Figure 8. Visualisation of heat transfer through IR sensing.

The performance of the prototype was monitored in realtime through digital temperature sensors and infrared emissions. Real time plots of DS18B20 temperature sensors in a prototype incubator show heating from room temperature to a set point of 60oC in two minutes. The microreactor could accurately maintain temperatures to within a degree at set points from 37o to 60o Celsius. This has been an encouraging start to the project, given the cost of the components, and with many avenues still to explore for improvement for efficiency, speed, cycling and real-time measurement of reactions in situ. A manufactured microreactor of this type would be cheap and find wide use for diagnostics, education and science, especially in low resource settings.

Further information: Test-rig assembly - https://www.hackster.io/jim-haseloff/programmable-test-rig-d7df62 Prototype microreactor - https://www.hackster.io/jim-haseloff/airloop-i-5d2a72 Biomaker - https://www.biomaker.org Biomaker Expert Group - contact Jim Haseloff at jh295@cam.ac.uk if you are interested in getting involved. All welcome!

Synthetic biology, coronavirus and me.

In common with most other people, I first became aware of the new coronavirus, SARS-CoV-2 (the causative agent of CoVID-19), through news reports from Wuhan in Hubei province, China in January 2020. At the time, it looked like this might be a re-run of the SARS epidemic from 2003, which was limited to China and neighbouring countries.

However, as the disease seemed to be spreading more quickly than SARS, I was asked to participate in a number of radio and TV interviews, wearing my virologist hat, starting on the 10th Feb. These early broadcasts mainly concerned a description of the virus and its symptoms and discussions of the containment measures being enforced in Wuhan and surrounding districts without much concern about the UK’s potential situation. This changed dramatically in early March when it became obvious that the disease had established itself in Italy and was rapidly moving throughout Europe.

Professor George Lomonossoff

The interviews became far more frequent and sombre in tone and started including questions about testing, social distancing, possible vaccines etc. I hope I was able to contribute useful information – it has certainly been a very interesting experience for me! I’m pleased to see that such concepts as PCR, antibody testing and R number are now in common parlance. A particular point I was able to stress was how quickly modern synthetic biology had enabled candidate vaccines to be produced, obviating the need to culture the infectious virus. However, demonstrating efficacy will take a bit longer.

Well, so much for my media work – what about a scientific contribution? Almost 20 years, I was involved in a project in the collaboration with the Pirbright Institute to develop qPCR controls to combat the 2001 outbreak of foot-and-mouth disease virus (FMDV). These were based on encapsidating FMDV sequences within cowpea mosaic virus (CPMV) particles Encapsidated mimic technology; N-Cap). This work, originally based on infectious CPMV, was successful and was applied to other veterinary diseases in collaboration the Veterinary Laboratories Agency (VLA; now called APHA).

In the past 2 years, Hadrien Peyret, a senior post-doc in my lab, has been working with Leaf Expression Systems and Meridian Biosciences to further develop the N-Cap system, using modern synthetic biology methods, as a source of PCR controls. As a result, we were in an ideal position to rapidly produce appropriate material for incorporation into qPCR-based testing kits for CoVID-19. Hadrien first discussed this on 17th March, the appropriate sequences were ordered on the 19th and arrived at JIC on the 30th.

Hadrien infiltrated plants 6th April, I harvested them on the 13th and by the 16th had purified the particles (yes folks, I really put my lab coat on and ran centrifuges!). The material is currently being evaluated by LES and St. George’s Hospital, London.

In addition to the qPCR controls my group has also been involved in expressing the structural proteins from SARS-CoV-2 in plants, with the aim of providing information to aid the development antibody tests and candidate vaccines. We were very fortunate that Jae-Wan Jung, a sabbatical visitor from S. Korea, has been working on the expression of the equivalent proteins of a related veterinary virus, porcine epidemic diarrhoea virus (PEDV). This gave us a head start and he agreed to shift the focus of his work to SARS-CoV-2.

To enable this, we applied for 12 months funding from the JIC Innovation Fund on the 30th March and received a positive response on 2nd April – not a bad turn-around and many thanks to all concerned. All the necessary DNA sequences have now arrived on-site, so it’s full steam ahead with expression studies.

That’s about it as of 1st May – stay safe.

Written by OpenPlan PI Professor George Lomonossoff

Bio-foundries in the Age of Pandemics

Biofoundries are integrated platforms for the rapid design, construction, and testing of genetically reprogrammed organisms for biotechnology applications and research. In practice, their application to biotechnology and genetic engineering lies mainly in the expertise of the scientists that work within them. Physically, Biofoundries are suites of laboratory automation equipment that could, conceivably, be applied to numerous scientific workflows. For example, the Earlham Biofoundy has five main automated platforms: an automated cryostorage system that will deliver a 96 well plate populated with a user-specified array of any of the 80K tubes in its store; a liquid handling system that moves nanolitre droplets of reagents (or cells) using acoustic energy; two highly flexible multi-station micro-scale liquid handling systems, and an automated microfermentation platform for optimising, monitoring and sampling microbial cultures. Similar equipment can be found in labs that conduct high-throughput genotyping, or in pathology labs, including diagnostic facilities.

Automation Platforms at the Earlham BioFoundry

Last year, the Global Biofoundry Alliance (GBA) was established to coordinate activities across the Biofoundries that are popping up across the world. While there are similarities in the goals of Foundries and, to an extent, their facilities, the projects and expertise varies between Foundries. Across the Biofoundries a wide array of molecular biology and biochemical workflows have been miniaturised, scaled and automated. In addition, numerous software packages have been developed to aid the design and analysis of experiments.

The Logo and Objectives of the Global Biofoundry Alliance

Along with specialised workflows for specific projects, most Biofoundries have miniaturised and automated at least one type of parallel DNA assembly reaction (e.g. Golden Gate and Gibson Assembly) together with several other core molecular biology workflows such as DNA/RNA purification, PCR set-up etc. All Foundries are able to perform at least hundreds of such reactions each week, and some have projects that require throughputs of thousands, or even tens of thousands.

As SARS-CoV-2, the virus strain that causes COVID-19, began to spread across the globe, it became clear that the pathology labs in many countries did not have the necessary capacity to meet demand. Further, it became apparent that the global demands for reagents required for testing, particularly for specific brands of reagents, were outstripping supply.  While a few Biofoundries with existing expertise or projects in diagnostics and infectious diseases have continued to work, unfortunately most Biofoundries have been shut down as they are not certified as diagnostics laboratories, and thus many automation platforms are sitting unused as Biofoundry staff moved to working from home.

However, the scientific community has responded astoundingly quickly with several inventive detection protocols being developed and published. It has also responded to support the infrastructures already approved for diagnostics. In Norwich, movable liquid-handling platforms were relocated from the Earlham Institute to the Norfolk and Norwich University Hospital to enable automation of the approved RNA-extraction protocol. At the time of writing, additional automation from the Earlham Biofoundry is being scoped for deactivation of virus on swabs and the preparation of reagents for the approved qPCR tests. The team at the London DNA Foundry have also relocated several automation platforms to their local hospitals to increase capacity in the capital. These automation platforms also require the presence of automation specialists to program and validate the workflows and to train new operators.

As pleased as we are to offer these small measures of assistance, there is a degree of frustration that the Biofoundries, filled with appropriate equipment for high throughput testing, are not enabled to do more. Interestingly, one of the exercises planned for the next meeting of the Global BioFoundries Alliance (previously planned for May 2020 in Canada), was to agree a series of activities that Biofoundries could undertake to address a Global Challenge such as emerging diseases. Sub-groups working on software, metrology and the development of the challenge were formed last year and had already began to formulate ideas. 

Since the pandemic has taken hold, we have continued these discussions with the main question being "Can Biofoundries develop generalised workflows for biosecurity threats, and, if so, what needs to be done?"

Interestingly, the many discussions all seemed to boil back down to the same aims that were initially described as the core business of the GBA:

·        To test the robustness of protocols across different platforms and locations

·        To establish infrastructures and a community of practice that would enable the rapid transfer and benchmarking of protocols on new platforms and in new locations

·        To establish and scale-up low-cost protocols, including those that can be performed on low-cost platforms

To these, Covid-19 has added:

·        The need to validate protocols using locally-manufactured reagents, which obviously includes the ability to make and validate the quality of key reagents such as reverse-transcriptases

·        A greater focus on the development of workflows on deployable automation platforms that can be relocated to the point-of-of need (e.g. approved diagnostic laboratories)

It is still unclear how much these current efforts will impact the current pandemic. But, this pandemic is highlighting some of the weak links in biosecurity and preparedness, showing us where we can and should focus our efforts to be able to deal with future challenges.

Written by Dr Nicola Patron, OpenPlant project leader.

Iceni Dianostics, an OpenPlant spinoff, contributes to the fight against COVID19.

In December 2019 the World Health Organisation received the first reports of a previously unknown virus, Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2).

Professor Rob Field, OpenPlant PI and Iceni Diagnostics cofounder.

Coronaviruses are a large group of viruses that are known to infect both humans and animals and cause a range of respiratory illnesses. The main symptoms of the virus outbreak, known as COVID19, include a fever and a continuous cough but in severe cases the disease can cause pneumonia and bronchitis which can cause death in some cases. 

The COVID19 outbreak has seemingly brought the world to its knees with many countries enforcing lockdown measures to reduce transmission rates and with over 32,000 deaths reported in the UK alone.

The scientific community has been fast to react to the novel outbreak, with the virus genome being discovered and published within the first four weeks of its detection. Now scientists across the globe are working to develop vaccines and diagnostic tools that help detect the virus.

With vaccine production and mass manufacture being lengthy and costly, many scientists are turning to developing diagnostic tests that can help in the bid to slow the transmission rate and spread of the disease at the point of care.

Many of the current COVID19 tests are largely based on PCR (polymerase chain reaction) that requires laboratory setting for analysis and knowledge of the virus’s genetic code.

However, Iceni Diagnostics, an OpenPlant spin off company based in Norwich are developing a new approach that identifies the virus using sugar (glycon) recognition instead of the viral genetic code, which can mutate.

Iceni Diagnostics existing prototype for influenza can detect the virus in less than 20 minutes, OpenPlant PI Professor Rob Field and his team are now working on adapting this test to detect the COVID19 virus.

The innovative handheld device requires a sample of saliva and uses lateral flow to give a simple positive or negative result. An important design feature is that it will require no training to operate allowing it to be used in any location by any individual.

To find out more about Iceni Diagnostics and their efforts to combat COVID19 visit their website here.

Digital Workshop: No-Code Programming for Biology

Sadly, in order to protect our community, the popular No-Code Programming for Biology workshop due to be held on 23rd-24th March was cancelled. However, we’re pleased to announce that the materials for this workshop are now being made available online.

This course was designed to introduce biologists to the basics of building custom instrumentation to assist with experiments in the lab and field. Part of the long-running Biomaker project, the course covers the fundamentals of using Arduino-based microcontrollers, sensor electronics, displays and actuators, as well as use of the visual programming language XOD. By combining these skills participants can learn to build a variety of instruments that are useful for measuring and controlling biological systems.

Making your own devices for use in biological research can be advantageous in many ways – components are often very cheap compared to commercially available options; there is a well-established community of experienced scientists focused on collaboration and open sharing of designs; and the opportunities for customisation and experiment-specific adjustments are endless. However, there are also hurdles for those looking to build custom instruments. Without a background in electronics or engineering it can be hard to know where to start. In addition, many systems require the maker to be able to understand and write code in languages such as CC+, which take time to learn and contain complex syntax.

Biomaker and the No-Code Programming for Biology workshop aim to break down some of these barriers. The course provides biologists with a hardware starter kit, a series of simple tutorials to get started using hardware and electronics, and an introduction to the visual programming language XOD. Instead of having to write code to control your instruments, XOD uses ‘nodes’ to represent different hardware and functions. Connecting these nodes in different combinations allows you to control your hardware and customise your instrumentation.

Previously, teams working on Biomaker projects have used these concepts for a wide variety of applications, including instrumentation, microscopy, microfluidics, 3D printing, biomedical devices, DNA design, plant sciences and outreach and public engagement. The No-Code programming for Biology workshop builds on what we have learned from running Biomaker projects and provides biologists with the necessary skills to start building their own devices and advance their research with inexpensive custom instrumentation.

If you would like to get involved with the No-Code Programming for Biology course the introduction, and first set of tutorials are now available on the Biomaker website. A number of hardware starter kits may be available upon request, although this cannot be guaranteed.

More information about the Biomaker project can be found on the Biomaker website. For questions and enquires please contact the Cambridge SynBio IRC and OpenPlant Coordinator Steph Norwood at coordinator@synbio.cam.ac.uk.