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Journal articleCrone MA, Priestman M, Ciechonska M, et al., 2020,
A role for Biofoundries in rapid development and validation of automated SARS-CoV-2 clinical diagnostics (vol 11, 4464, 2020), NATURE COMMUNICATIONS, Vol: 11, ISSN: 2041-1723
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- Citations: 1
Journal articleCrone M, Priestman M, Ciechonska M, et al., 2020,
A role for Biofoundries in rapid development and validation of automated SARS-CoV-2 clinical diagnostics, Nature Communications, Vol: 11, Pages: 1-11, ISSN: 2041-1723
The SARS-CoV-2 pandemic has shown how a rapid rise in demand for patient and community sample testing can quickly overwhelm testing capability globally. With most diagnostic infrastructure dependent on specialized instruments, their exclusive reagent supplies quickly become bottlenecks, creating an urgent need for approaches to boost testing capacity. We address this challenge by refocusing the London Biofoundry onto the development of alternative testing pipelines. Here, we present a reagent-agnostic automated SARS-CoV-2 testing platform that can be quickly deployed and scaled. Using an in-house-generated, open-source, MS2-virus-like particle (VLP) SARS-CoV-2 standard, we validate RNA extraction and RT-qPCR workflows as well as two detection assays based on CRISPR-Cas13a and RT-loop-mediated isothermal amplification (RT-LAMP). In collaboration with an NHS diagnostic testing lab, we report the performance of the overall workflow and detection of SARS-CoV-2 in patient samples using RT-qPCR, CRISPR-Cas13a, and RT-LAMP. The validated RNA extraction and RT-qPCR platform has been installed in NHS diagnostic labs, increasing testing capacity by 1000 samples per day.
Journal articleGraham N, Junghans C, Downes R, et al., 2020,
SARS-CoV-2 infection, clinical features and outcome of COVID-19 in United Kingdom nursing homes, Journal of Infection, Vol: 81, Pages: 411-419, ISSN: 0163-4453
OBJECTIVES: To understand SARS-Co-V-2 infection and transmission in UK nursing homes in order to develop preventive strategies for protecting the frail elderly residents. METHODS: An outbreak investigation involving 394 residents and 70 staff, was carried out in 4 nursing homes affected by COVID-19 outbreaks in central London. Two point-prevalence surveys were performed one week apart where residents underwent SARS-CoV-2 testing and had relevant symptoms documented. Asymptomatic staff from three of the four homes were also offered SARS-CoV-2 testing. RESULTS: Overall, 26% (95% CI 22 to 31) of residents died over the two-month period. All-cause mortality increased by 203% (95% CI 70 to 336) compared with previous years. Systematic testing identified 40% (95% CI 35 to 46) of residents as positive for SARS-CoV-2, and of these 43% (95% CI 34 to 52) were asymptomatic and 18% (95% CI 11 to 24) had only atypical symptoms; 4% (95% CI -1 to 9) of asymptomatic staff also tested positive. CONCLUSIONS: The SARS-CoV-2 outbreak in four UK nursing homes was associated with very high infection and mortality rates. Many residents developed either atypical or no discernible symptoms. A number of asymptomatic staff members also tested positive, suggesting a role for regular screening of both residents and staff in mitigating future outbreaks.
Journal articleRamlaul K, Palmer C, Nakane T, et al., 2020,
Mitigating local over-fitting during single particle reconstruction with SIDESPLITTER, Journal of Structural Biology, Vol: 211, Pages: 1-9, ISSN: 1047-8477
Single particle analysis has become a key structural biology technique. Experimental images are extremely noisy, and during iterative refinement it is possible to stably incorporate noise into the reconstruction. Such “over-fitting” can lead to misinterpretation of the structure and flawed biological results. Several strategies are routinely used to prevent over-fitting, the most common being independent refinement of two sides of a split dataset. In this study, we show that over-fitting remains an issue within regions of low local signal-to-noise, despite independent refinement of half datasets. We propose a modification of the refinement process through the application of a local signal-to-noise filter: SIDESPLITTER. We show that our approach can reduce over-fitting for both idealised and experimental data while maintaining independence between the two sides of a split refinement. SIDESPLITTER refinement leads to improved density, and can also lead to improvement of the final resolution in extreme cases where datasets are prone to severe over-fitting, such as small membrane proteins.
Journal articleKelwick R, Webb A, Freemont P, 2020,
Biological materials: the next frontier for cell-free synthetic biology, Frontiers in Bioengineering and Biotechnology, Vol: 8, ISSN: 2296-4185
Advancements in cell-free synthetic biology are enabling innovations in sustainable biomanufacturing, that may ultimately shift the global manufacturing paradigm toward localized and ecologically harmonized production processes. Cell-free synthetic biology strategies have been developed for the bioproduction of fine chemicals, biofuels and biological materials. Cell-free workflows typically utilize combinations of purified enzymes, cell extracts for biotransformation or cell-free protein synthesis reactions, to assemble and characterize biosynthetic pathways. Importantly, cell-free reactions can combine the advantages of chemical engineering with metabolic engineering, through the direct addition of co-factors, substrates and chemicals –including those that are cytotoxic. Cell-free synthetic biology is also amenable to automatable design cycles through which an array of biological materials and their underpinning biosynthetic pathways can be tested and optimized in parallel. Whilst challenges still remain, recent convergences between the materials sciences and these advancements in cell-free synthetic biology enable new frontiers for materials research.
Journal articleBeal J, Goñi-Moreno A, Myers C, et al., 2020,
The long journey towards standards for engineering biosystems: Are the Molecular Biology and the Biotech communities ready to standardise?, EMBO Reports, Vol: 21, Pages: 1-5, ISSN: 1469-221X
Synthetic biology needs to adopt sound scientific and industry-like standards in order to achieve its ambitious goals of efficient and accurate engineering of biological systems.
Journal articleCrone MA, Priestman M, Ciechonska M, et al., 2020,
A new role for Biofoundries in rapid prototyping, development, and validation of automated clinical diagnostic tests for SARS-CoV-2
<jats:title>Abstract</jats:title><jats:p>The SARS-CoV-2 pandemic has shown how the rapid rise in demand for patient and community sample testing, required for tracing and containing a highly infectious disease, has quickly overwhelmed testing capability globally. With most diagnostic infrastructure dependent on specialised instruments, their exclusive reagent supplies quickly become bottlenecks in times of peak demand, creating an urgent need for novel approaches to boost testing capacity. We address this challenge by refocusing the full synthetic biology stack available at the London Biofoundry onto the development of alternative patient sample testing pipelines. We present a reagent-agnostic automated SARS-CoV-2 testing platform that can be quickly deployed and scaled, and that accepts a diverse range of reagents. Using an in-house-generated, open-source, MS2-virus-like-particle-SARS-CoV-2 standard, we validate RNA extraction and RT-qPCR workflows as well as two novel detection assays based on CRISPR-Cas and Loop-mediated isothermal Amplification (LAMP) approaches. In collaboration with an NHS diagnostic testing lab, we report the performance of the overall workflow and benchmark SARS-CoV-2 detection in patient samples via RT-qPCR, CRISPR-Cas, and LAMP against clinical test sets. The validated RNA extraction and RT-qPCR platform has been installed in NHS diagnostic labs and now contributes to increased patient sample processing in the UK while we continue to refine and develop novel high-throughput diagnostic methods. Finally, our workflows and protocols can be quickly implemented and adapted by members of the Global Biofoundry Alliance and the wider scientific and medical diagnostics community.</jats:p>
Journal articleWilkinson MD, Lai H-E, Freemont PS, et al., 2020,
A biosynthetic platform for antimalarial drug discovery, Antimicrobial Agents and Chemotherapy, Vol: 64, Pages: 1-9, ISSN: 0066-4804
Advances in synthetic biology have enabled production of a variety of compounds using bacteria as a vehicle for complex compound biosynthesis. Violacein, a naturally occurring indole pigment with antibiotic properties, can be biosynthetically engineered in Escherichia coli expressing its non-native synthesis pathway. To explore whether this synthetic biosynthesis platform could be used for drug discovery, here we have screened bacterially-derived violacein against the main causative agent of human malaria, Plasmodium falciparum. We show the antiparasitic activity of bacterially-derived violacein against the P. falciparum 3D7 laboratory reference strain as well as drug-sensitive and resistant patient isolates, confirming the potential utility of this drug as an antimalarial. We then screen a biosynthetic series of violacein derivatives against P. falciparum growth. The demonstrated varied activity of each derivative against asexual parasite growth points to potential for further development of violacein as an antimalarial. Towards defining its mode of action, we show that biosynthetic violacein affects the parasite actin cytoskeleton, resulting in an accumulation of actin signal that is independent of actin polymerization. This activity points to a target that modulates actin behaviour in the cell either in terms of its regulation or its folding. More broadly, our data show that bacterial synthetic biosynthesis could become a suitable platform for antimalarial drug discovery with potential applications in future high-throughput drug screening with otherwise chemically-intractable natural products.
Journal articleMoore SJ, Lai H-E, Kelwick RJR, et al., 2020,
Correction to EcoFlex: a multifunctional MoClo kit for E. coli synthetic biology., ACS Synthetic Biology, ISSN: 2161-5063
It has been brought to our attention that the original article contains a typographical error within Figure 1B, part ii. One of the 4-bp overhangs reads “GGAC” and should instead be “GTAC”, as is consistent throughout the original manuscript and deposited AddGene sequences.
Journal articleWilliams R, Yates L, Zhang X, 2020,
Structures and regulations of ATM and ATR, master kinases in genome integrity, Current Opinion in Structural Biology, Vol: 61, Pages: 98-105, ISSN: 0959-440X
Homologous recombination (HR) is a faithful repair mechanism for double stranded DNA breaks. Two highly homologous master kinases, the tumour suppressors ATM and ATR (Tel1 and Mec1 in yeast), coordinate cell cycle progression with repair during HR. Despite their importance, our molecular understanding of these apical coordinators has been limited, in part due to their large sizes. With the recent development in cryo-electron microscopy, significant advances have been made in structural characterisation of these proteins in the last two years. These structures, combined with new biochemical studies, now provide a more detailed understanding of how a low basal activity is maintained and how activation may occur. In this review, we summarize recent advances in the structural and molecular understanding of these key components in HR, compare the common and distinct features of these kinases and suggest aspects of structural components that are likely to be involved in regulating its activity.
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