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Synthetic Biology underpins advances in the bioeconomy

Biological systems - including the simplest cells - exhibit a broad range of functions to thrive in their environment. Research in the Imperial College Centre for Synthetic Biology is focused on the possibility of engineering the underlying biochemical processes to solve many of the challenges facing society, from healthcare to sustainable energy. In particular, we model, analyse, design and build biological and biochemical systems in living cells and/or in cell extracts, both exploring and enhancing the engineering potential of biology. 

As part of our research we develop novel methods to accelerate the celebrated Design-Build-Test-Learn synthetic biology cycle. As such research in the Centre for Synthetic Biology highly multi- and interdisciplinary covering computational modelling and machine learning approaches; automated platform development and genetic circuit engineering ; multi-cellular and multi-organismal interactions, including gene drive and genome engineering; metabolic engineering; in vitro/cell-free synthetic biology; engineered phages and directed evolution; and biomimetics, biomaterials and biological engineering.


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  • Journal article
    Stach L, Morgan RM, Makhlouf L, Douangamath A, von Delft F, Zhang X, Freemont PSet al., 2020,

    Crystal structure of the catalytic D2 domain of the AAA+ ATPase p97 reveals a putative helical split-washer-type mechanism for substrate unfolding

    , FEBS Letters, Vol: 594, Pages: 933-943, ISSN: 0014-5793

    Several pathologies have been associated with the AAA+ ATPase p97, an enzyme essential to protein homeostasis. Heterozygous polymorphisms in p97 have been shown to cause neurological disease, while elevated proteotoxic stress in tumours has made p97 an attractive cancer chemotherapy target. The cellular processes reliant on p97 are well described. High‐resolution structural models of its catalytic D2 domain, however, have proved elusive, as has the mechanism by which p97 converts the energy from ATP hydrolysis into mechanical force to unfold protein substrates. Here, we describe the high‐resolution structure of the p97 D2 ATPase domain. This crystal system constitutes a valuable tool for p97 inhibitor development and identifies a potentially druggable pocket in the D2 domain. In addition, its P61 symmetry suggests a mechanism for substrate unfolding by p97.

  • Journal article
    Riangrungroj P, Polizzi KM, 2020,

    BeQuIK (Biosensor Engineered Quorum Induced Killing): designer bacteria for destroying recalcitrant biofilms.

    , Microbial Biotechnology, Vol: 13, Pages: 311-314, ISSN: 1751-7915

    This opinion piece describes a new design for the remediation of recalcitrant biofilms. It builds on previous work to develop engineered E. coli that recognize quorum sensing signals from pathogens in a biofilm and secrete toxins in response. To solve the challenge of dilute signalling molecules, we propose to use nanobodies and enzymes displayed on the surface of the cells to localize them to the biofilm and degrade the extracellular polymeric substances, thus creating a solution with better 'seek and destroy' capabilities.

  • Journal article
    Spice AJ, Aw R, Bracewell DG, Polizzi KMet al., 2020,

    Synthesis and assembly of Hepatitis B virus-like particles in a Pichia pastoris cell-free system

    , Frontiers in Bioengineering and Biotechnology, Vol: 8, ISSN: 2296-4185

    Virus-like particles (VLPs) are supramolecular protein assemblies with the potential for unique and exciting applications in synthetic biology and medicine. Despite the attention VLPs have gained thus far, considerable limitations still persist in their production. Poorly scalable manufacturing technologies and inconsistent product architectures continue to restrict the full potential of VLPs. Cell-free protein synthesis (CFPS) offers an alternative approach to VLP production and has already proven to be successful, albeit using extracts from a limited number of organisms. Using a recently developed Pichia pastoris-based CFPS system, we have demonstrated the production of the model Hepatitis B core antigen VLP as a proof-of-concept. The VLPs produced in the CFPS system were found to have comparable characteristics to those previously produced in vivo and in vitro. Additionally, we have developed a facile and rapid synthesis, assembly and purification methodology that could be applied as a rapid prototyping platform for vaccine development or synthetic biology applications. Overall the CFPS methodology allows far greater throughput, which will expedite the screening of optimal assembly conditions for more robust and stable VLPs. This approach could therefore support the characterization of larger sample sets to improve vaccine development efficiency.

  • Journal article
    Jacobsen IH, Ledesma-Amaro R, Martinez JL, 2020,

    Recombinant β-carotene production by yarrowia lipolytica - assessing the potential of micro-scale fermentation analysis in cell factory design and bioreaction optimization

    , Frontiers in Bioengineering and Biotechnology, Vol: 8, ISSN: 2296-4185

    The production of β-carotene has become increasingly interesting within the biotechnological industry due to a rising demand for safer and more natural colorants, nutritional supplements, and antioxidants. A recent study has described the potential of Yarrowia lipolytica as a β-carotene-producing cell factory, reporting the highest titer of recombinant β-carotene produced to date. Finding the best conditions to maximize production and scaling up the process to full scale, a costly and time-consuming process, it is often a bottleneck in biotechnology. In this work, we explored the benefits of using micro-fermentation equipment to significantly reduce the time spent on design and optimization of bioreaction conditions, especially in the early stages of process development. In this proof-of-concept study, a β-carotene producing Y. lipolytica strain was tested in micro-fermentations partly to assess the robustness of the cell factory design and partly to perform media optimization. The medium optimization led us to an improvement of up to 50% in the yield of β-carotene production in the best of the conditions. Overall, the micro-fermentation system had a high degree of reliability in all tests.

  • Journal article
    Gowers G, Chee S, Bell D, Suckling L, Kern M, Tew D, McClymont D, Ellis Tet al., 2020,

    Improved betulinic acid biosynthesis using synthetic yeast chromosome recombination and semi-automated rapid LC-MS screening

    , Nature Communications, Vol: 11, ISSN: 2041-1723

    Synthetic biology, genome engineering and directed evolution offer innumerable tools to expedite engineering of strains for optimising biosynthetic pathways. One of the most radical is SCRaMbLE, a system of inducible in vivo deletion and rearrangement of synthetic yeast chromosomes, diversifying the genotype of millions of Saccharomyces cerevisiae cells in hours. SCRaMbLE can yield strains with improved biosynthetic phenotypes but is limited by screening capabilities. To address this bottleneck, we combine automated sample preparation, an ultra-fast 84-second LC-MS method, and barcoded nanopore sequencing to rapidly isolate and characterise the best performing strains. Here, we use SCRaMbLE to optimise yeast strains engineered to produce the triterpenoid betulinic acid. Our semi-automated workflow screens 1,000 colonies, identifying and sequencing 12 strains with between 2- to 7-fold improvement in betulinic acid titre. The broad applicability of this workflow to rapidly isolate improved strains from a variant library makes this a valuable tool for biotechnology.

  • Journal article
    Wu Y, Chen T, Liu Y, Tian R, Lv X, Li J, Du G, Chen J, Ledesma-Amaro R, Liu Let al., 2020,

    Design of a programmable biosensor-CRISPRi genetic circuits for dynamic and autonomous dual-control of metabolic flux in Bacillus subtilis

    , Nucleic Acids Research, Vol: 48, Pages: 996-1009, ISSN: 0305-1048

    Dynamic regulation is an effective strategy for fine-tuning metabolic pathways in order to maximize target product synthesis. However, achieving dynamic and autonomous up- and down-regulation of the metabolic modules of interest simultaneously, still remains a great challenge. In this work, we created an autonomous dual-control (ADC) system, by combining CRISPRi-based NOT gates with novel biosensors of a key metabolite in the pathway of interest. By sensing the levels of the intermediate glucosamine-6-phosphate (GlcN6P) and self-adjusting the expression levels of the target genes accordingly with the GlcN6P biosensor and ADC system enabled feedback circuits, the metabolic flux towards the production of the high value nutraceutical N-acetylglucosamine (GlcNAc) could be balanced and optimized in Bacillus subtilis. As a result, the GlcNAc titer in a 15-l fed-batch bioreactor increased from 59.9 g/l to 97.1 g/l with acetoin production and 81.7 g/l to 131.6 g/l without acetoin production, indicating the robustness and stability of the synthetic circuits in a large bioreactor system. Remarkably, this self-regulatory methodology does not require any external level of control such as the use of inducer molecules or switching fermentation/environmental conditions. Moreover, the proposed programmable genetic circuits may be expanded to engineer other microbial cells and metabolic pathways.

  • Journal article
    Wen Z, Ledesma-Amaro R, Lu M, Jin M, Yang Set al., 2020,

    Metabolic engineering of clostridium cellulovorans to improve butanol production by consolidated bioprocessing.

    , ACS Synthetic Biology, Vol: 9, Pages: 304-315, ISSN: 2161-5063

    Clostridium cellulovorans DSM 743B can produce butyrate when grown on lignocellulose, but it can hardly synthesize butanol. In a previous study, C. cellulovorans was successfully engineered to switch the metabolism from butyryl-CoA to butanol by overexpressing an alcohol aldehyde dehydrogenase gene adhE1 from Clostridium acetobutylicum ATCC 824; however, its full potential in butanol production is still unexplored. In the study, a metabolic engineering approach based on a push-pull strategy was developed to further enhance cellulosic butanol production. In order to accomplish this, the carbon flux from acetyl-CoA to butyryl-CoA was pulled by overexpressing a trans-enoyl-coenzyme A reductase gene (ter), which can irreversibly catalyze crotonyl-CoA to butyryl-CoA. Then an acid reassimilation pathway uncoupled with acetone production was introduced to redirect the carbon flow from butyrate and acetate toward butyryl-CoA. Finally, xylose metabolism engineering was implemented by inactivating xylR (Clocel_0594) and araR (Clocel_1253), as well as overexpressing xylT (CA_C1345), which is expected to supply additional carbon and reducing power for CoA and butanol synthesis pathways. The final engineered strain produced 4.96 g/L of n-butanol from alkali extracted corn cobs (AECC), increasing by 235-fold compared to that of the wild type. It serves as a promising butanol producer by consolidated bioprocessing.

  • Journal article
    Park Y-K, Ledesma-Amaro R, Nicaud J-M, 2020,

    De novo biosynthesis of odd-chain fatty acids in yarrowia lipolytica enabled by modular pathway engineering

    , Frontiers in Bioengineering and Biotechnology, Vol: 7, Pages: 1-11, ISSN: 2296-4185

    Microbial oils are regarded as promising alternatives to fossil fuels as concerns over environmental issues and energy production systems continue to mount. Odd-chain fatty acids (FAs) are a type of valuable lipid with various applications: they can serve as biomarkers, intermediates in the production of flavor and fragrance compounds, fuels, and plasticizers. Microorganisms naturally produce FAs, but such FAs are primarily even-chain; only negligible amounts of odd-chain FAs are generated. As a result, studies using microorganisms to produce odd-chain FAs have had limited success. Here, our objective was to biosynthesize odd-chain FAs de novo in Yarrowia lipolytica using inexpensive carbon sources, namely glucose, without any propionate supplementation. To achieve this goal, we constructed a modular metabolic pathway containing seven genes. In the engineered strain expressing this pathway, the percentage of odd-chain FAs out of total FAs was higher than in the control strain (3.86 vs. 0.84%). When this pathway was transferred into an obese strain, which had been engineered to accumulate large amounts of lipids, odd-chain fatty acid production was 7.2 times greater than in the control (0.05 vs. 0.36 g/L). This study shows that metabolic engineering research is making progress toward obtaining efficient cell factories that produce odd-chain FAs.

  • Journal article
    Sattayawat P, Yunus IS, Jones PR, 2020,

    Bioderivatization as a concept for renewable production of chemicals that are toxic or poorly soluble in the liquid phase

    , Proceedings of the National Academy of Sciences of USA, Vol: 117, Pages: 1404-1413, ISSN: 0027-8424

    Bio-based production technologies may complement or replace petroleum-based production of chemicals, but they face a number of technical challenges, including product toxicity and/or water insolubility. Plants and microorganisms naturally biosynthesize chemicals that often are converted into derivatives with reduced toxicity or enhanced solubility. Inspired by this principle, we propose a bioderivatization strategy for biotechnological chemicals production, defined as purposeful biochemical derivatization of intended target molecules. As proof of principle, the effects of hydrophobic (e.g., esterification) and hydrophilic (e.g., glycosylation) bioderivatization strategies on the biosynthesis of a relatively toxic and poorly soluble chemical, 1-octanol, were evaluated in Escherichia coli and Synechocystis sp. PCC 6803. The 1-octanol pathway was first optimized to reach product titers at which the host displayed symptoms of toxicity. Solvent overlay used to capture volatile products partially masked product toxicity. Regardless of whether solvent overlay was used, most strains with bioderivatization had a higher molar product titer and product yield, as well as improved cellular growth and glucose consumption, compared with strains without bioderivatization. The positive effect on bioproduction was observed with both the hydrophobic and hydrophilic strategies. Interestingly, in several combinations of genotype/induction strength, bioderivatization had a positive effect on productivity without any apparent effect on growth. We attribute this to enhanced product solubility in the aqueous or solvent fraction of the bioreactor liquid phase (depending on the derivative and medium used), with consequent enhanced product removal. Overall, under most conditions, a benefit of bioproduction was observed, and the bioderivatization strategy could be considered for other similar chemicals as well.

  • Journal article
    Yunus IS, Palma A, Trudeau DL, Tawfik DS, Jones PRet al., 2020,

    Methanol-free biosynthesis of fatty acid methyl ester (FAME) in Synechocystis sp. PCC 6803

    , METABOLIC ENGINEERING, Vol: 57, Pages: 217-227, ISSN: 1096-7176
  • Journal article
    Kopniczky MB, Canavan C, McClymont DW, Crone MA, Suckling L, Goetzmann B, Siciliano V, MacDonald JT, Jensen K, Freemont PSet al., 2020,

    Cell-free protein synthesis as a prototyping platform for mammalian synthetic biology

    , ACS Synthetic Biology, Vol: 9, Pages: 144-156, ISSN: 2161-5063

    The field of mammalian synthetic biology is expanding quickly, and technologies for engineering large synthetic gene circuits are increasingly accessible. However, for mammalian cell engineering, traditional tissue culture methods are slow and cumbersome, and are not suited for high-throughput characterization measurements. Here we have utilized mammalian cell-free protein synthesis (CFPS) assays using HeLa cell extracts and liquid handling automation as an alternative to tissue culture and flow cytometry-based measurements. Our CFPS assays take a few hours, and we have established optimized protocols for small-volume reactions using automated acoustic liquid handling technology. As a proof-of-concept, we characterized diverse types of genetic regulation in CFPS, including T7 constitutive promoter variants, internal ribosomal entry sites (IRES) constitutive translation-initiation sequence variants, CRISPR/dCas9-mediated transcription repression, and L7Ae-mediated translation repression. Our data shows simple regulatory elements for use in mammalian cells can be quickly prototyped in a CFPS model system.

  • Journal article
    de Martín Garrido N, Crone MA, Ramlaul K, Simpson PA, Freemont PS, Aylett CHSet al., 2020,

    Bacteriophage MS2 displays unreported capsid variability assembling T = 4 and mixed capsids

    , Molecular Microbiology, Vol: 113, Pages: 143-152, ISSN: 0950-382X

    Bacteriophage MS2 is a positive-sense, single-stranded RNA virus encapsulated in an asymmetric T = 3 pseudo-icosahedral capsid. It infects Escherichia coli through the F-pilus, which it binds through a maturation protein incorporated into its capsid. Cryogenic electron microscopy has previously shown that its genome is highly ordered within virions, and that it regulates the assembly process of the capsid. In this study we have assembled recombinant MS2 capsids with non-genomic RNA containing the capsid incorporation sequence, and investigated the structures formed, revealing that T = 3, T = 4 and mixed capsids between these two triangulation numbers are generated, and resolving structures of T = 3 and T = 4 capsids to 4 Å and 6 Å respectively. We conclude that the basic MS2 capsid can form a mix of T = 3 and T = 4 structures, supporting a role for the ordered genome in favouring the formation of functional T = 3 virions.

  • Journal article
    Wen Z, Lu M, Ledesma-Amaro R, Li Q, Jin M, Yang Set al., 2020,

    TargeTron technology applicable in solventogenic clostridia: Revisiting 12 years’ advances

    , Biotechnology Journal, Vol: 15, Pages: 1-14, ISSN: 1860-6768

    Clostridium has great potential in industrial application and medical research. But low DNA repair capacity and plasmids transformation efficiency severely delayed development and application of genetic tools based on homologous recombination (HR). TargeTron is a gene editing technique dependent on the mobility of group II introns, rather than homologous recombination, which made it very suitable for gene disruption of Clostridium. The application of TargeTron technology in Clostridium was academically reported in 2007 and this tool has been introduced in various clostridia as it is easy to operate, time-saving, and reliable. TargeTron has made great progress in solventogenic Clostridium in the aspects of acetone-butanol-ethanol (ABE) fermentation pathway modification, important functional genes identification, and xylose metabolic pathway analysis & reconstruction. In the review, we revisited 12 years' advances of TargeTron technology applicable in solventogenic Clostridium, including its principle, technical characteristics, application and efforts to expand its capabilities, or to avoid potential drawbacks. Some other technologies as putative competitors or collaborators are also discussed. We believe that TargeTron combined with CRISPR/Cas-assisted gene/base editing and gene-expression regulation system will make a better future for clostridial genetic modification.

  • Book chapter
    Sootla A, Stan G-B, Ernst D, 2020,

    Solving Optimal Control Problems for Monotone Systems Using the Koopman Operator

    , Lecture Notes in Control and Information Sciences, Publisher: Springer International Publishing, Pages: 283-312, ISBN: 9783030357122

    © Springer Nature Switzerland AG 2020. Koopman operator theory offers numerous techniques for analysis and control of complex systems. In particular, in this chapter we will argue that for the problem of convergence to an equilibrium, the Koopman operator can be used to take advantage of the geometric properties of controlled systems, thus making the optimal solutions more transparent, and easier to analyze and implement. The motivation for the study of the convergence problem comes from biological applications, where easy-to-implement and easy-to-analyze solutions are of particular value. At the moment, theoretical results have been developed for a class of nonlinear systems called monotone systems. However, the core ideas presented here can be applied heuristically to non-monotone systems. Furthermore, the convergence problem can serve as a building block for solving other control problems such as switching between stable equilibria or inducing oscillations. These applications are illustrated in biologically inspired numerical examples.

  • Journal article
    Storch M, Dwijayanti A, Mallick H, Haines MC, Baldwin GSet al., 2020,

    BASIC: A Simple and Accurate Modular DNA Assembly Method.

    , Methods Mol Biol, Vol: 2205, Pages: 239-253

    Biopart Assembly Standard for Idempotent Cloning (BASIC) is a simple, robust, and highly accurate DNA assembly method, which provides 99% correct assemblies for a typical four-part assembly, enabling high efficiency cloning workflows (Storch et al., ACS Synth Biol, , 2015). BASIC employs standardised DNA linkers to combine bioparts, stored in the universal BASIC format. Once a new biopart is formatted into BASIC standard, defined by flanking 18 bp prefix and suffix sequences, it can be placed at any position and in any context within a designed BASIC assembly. This modularity of the BASIC approach is further enhanced by a range of functional linkers, including genetic elements like ribosomal binding sites (RBS) and peptide linkers. The method has a single tier format, whereby any BASIC assembly can create a new composite BASIC part in the same format used for the original parts; it can thus enter a subsequent BASIC assembly without the need for reformatting or changes to the workflow. This unique idempotent cloning mechanism allows for the assembly of constructs in multiple, conceptionally simple hierarchical rounds. Combined with its high accuracy and robustness, this makes BASIC a versatile assembly method for combinatorial and complex assemblies both at bench and biofoundry scale. The single universal storage format of BASIC parts enables compressed universal biopart libraries that promote sharing of parts and reproducible assembly strategies across labs, supporting efforts to improve reproducibility. In comparison with other DNA assembly standards and methods, BASIC offers a simple robust protocol, relies on a single tier format, provides for easy hierarchical assembly, and is highly accurate for up to seven parts per assembly round (Casini et al., Nat Rev Mol Cell Biol. , 2015).

  • Journal article
    Angrisano F, Sala K, Tapanelli S, Christophides G, Blagborough Aet al., 2019,

    Male-specific protein disulphide isomerase function is essential for plasmodium transmission and a vulnerable target for intervention

    , Scientific Reports, Vol: 9, ISSN: 2045-2322

    Inhibiting transmission of Plasmodium is an essential strategy in malaria eradication, and the biological process of gamete fusion during fertilization is a proven target for this approach. Lack of knowledge of the mechanisms underlying fertilization have been a hindrance in the development of transmission-blocking interventions. Here we describe a protein disulphide isomerase essential for malarial transmission (PDI-Trans/PBANKA_0820300) to the mosquito. We show that PDI-Trans activity is male-specific, surface-expressed, essential for fertilization/transmission, and exhibits disulphide isomerase activity which is up-regulated post-gamete activation. We demonstrate that PDI-Trans is a viable anti-malarial drug and vaccine target blocking malarial transmission with the use of PDI inhibitor bacitracin (98.21%/92.48% reduction in intensity/prevalence), and anti-PDI-Trans antibodies (66.22%/33.16% reduction in intensity/prevalence). To our knowledge, these results provide the first evidence that PDI function is essential for malarial transmission, and emphasize the potential of anti-PDI agents to act as anti-malarials, facilitating the future development of novel transmission-blocking interventions.

  • Journal article
    Jezierska S, Claus S, Ledesma-Amaro R, Van Bogaert Iet al., 2019,

    Redirecting the lipid metabolism of the yeast Starmerella bombicola from glycolipid to fatty acid production

    , Journal of Industrial Microbiology and Biotechnology, Vol: 46, Pages: 1697-1706, ISSN: 0169-4146

    Free fatty acids are basic oleochemicals implemented in a range of applications including surfactants, lubricants, paints, plastics, and cosmetics. Microbial fatty acid biosynthesis has gained much attention as it provides a sustainable alternative for petrol- and plant oil-derived chemicals. The yeast Starmerella bombicola is a microbial cell factory that naturally employs its powerful lipid metabolism for the production of the biodetergents sophorolipids (> 300 g/L). However, in this study we exploit the lipidic potential of S. bombicola and convert it from the glycolipid production platform into a free fatty acid cell factory. We used several metabolic engineering strategies to promote extracellular fatty acid accumulation which include blocking competing pathways (sophorolipid biosynthesis and β-oxidation) and preventing free fatty acid activation. The best producing mutant (Δcyp52m1Δfaa1Δmfe2) secreted 0.933 g/L (± 0.04) free fatty acids with a majority of C18:1 (43.8%) followed by C18:0 and C16:0 (40.0 and 13.2%, respectively). Interestingly, deletion of SbFaa1 in a strain still producing sophorolipids also resulted in 25% increased de novo sophorolipid synthesis (P = 0.0089) and when oil was supplemented to the same strain, a 50% increase in sophorolipid production was observed compared to the wild type (P = 0.03). We believe that our work is pivotal for the further development and exploration of S. bombicola as a platform for synthesis of environmentally friendly oleochemicals.

  • Journal article
    Scholes NS, Schnoerr D, Isalan M, Stumpf MPHet al., 2019,

    A Comprehensive Network Atlas Reveals That Turing Patterns Are Common but Not Robust

    , CELL SYSTEMS, Vol: 9, Pages: 515-517, ISSN: 2405-4712
  • Journal article
    Casula E, Traversari G, Fadda S, Klymenko O, Kontoravdi C, Cincotti Aet al., 2019,

    Modelling the osmotic behaviour of human mesenchymal stem cells

  • Journal article
    Gowers G-OF, Cameron SJS, Perdones-Montero A, Bell D, Chee SM, Kern M, Tew D, Ellis T, Takats Zet al., 2019,

    Off-colony screening of biosynthetic libraries by rapid laser-enabled mass spectrometry

    , ACS Synthetic Biology, Vol: 8, Pages: 2566-2575, ISSN: 2161-5063

    Leveraging advances in DNA synthesis and molecular cloning techniques, synthetic biology increasingly makes use of large construct libraries to explore large design spaces. For biosynthetic pathway engineering the ability to screen these libraries for a variety of metabolites of interest is essential. If the metabolite of interest or the metabolic phenotype is not easily measurable, screening soon becomes a major bottleneck involving time-consuming culturing, sample preparation, and extraction. To address this, we demonstrate the use of automated Laser-Assisted Rapid Evaporative Ionisation Mass Spectrometry (LA-REIMS) - a form of ambient laser desorption ionisation mass spectrometry - to perform rapid mass spectrometry analysis direct from agar plate yeast colonies without sample preparation or extraction. We use LA-REIMS to assess production levels of violacein and betulinic acid directly from yeast colonies at a rate of 6 colonies per minute. We then demonstrate the throughput enabled by LA-REIMS by screening over 450 yeast colonies in under 4 hours, while simultaneously generating recoverable glycerol stocks of each colony in real-time. This showcases LA-REIMS as a pre-screening tool to complement downstream quantification methods such as LCMS. Through pre-screening several hundred colonies with LA-REIMS, we successfully isolate and verify a strain with a 2.5-fold improvement in betulinic acid production. Finally, we show that LA-REIMS can detect 20 out of a panel of 27 diverse biological molecules, demonstrating the broad applicability of LA-REIMS to metabolite detection. The rapid and automated nature of LA-REIMS makes this a valuable new technology to complement existing screening technologies currently employed in academic and industrial workflows.

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