2023 PARTICIPATING UNIVERSITIES
Cas Catcher - MQ
Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 gene editing technology allows for precise genome editing within an organism through a cut site generated by the Cas9 enzyme. Successful transformation of the organism is reliant on endogenous genome repair machinery; mainly homology-directed repair (HDR). However, this technology is limited by high rates of Cas induced lethality due to low HDR efficiency, making this technology unfeasible in a wide range of organisms. We propose Cas Catcher will aid HDR and increase efficiency by adapting a system created by Andrew Hao and colleagues from Adelaide University (1). In which an orthogonal dCas dimer pair connected via a leucine zipper will hold the double stranded DNA break in physical proximity to assist genetic repair and incorporation of the novel gene. This project is a proof of concept on DNA looping’s capability to increase the yield of GFP knock-ins in Escherichia coli. Cas Catcher is our proposed solution to improve the toolbox capabilities of CRISPR-Cas9 genome engineering in non-model organisms traditionally restricted from CRISPR.
(1) Hao, N., Shearwin, K.E. and Dodd, I.B., 2017. Programmable DNA looping using engineered bivalent dCas9 complexes. Nature communications, 8(1), pp.1-12.
DeNovocastrians - UON
The DeNovocastrians aim to address the environmental contamination by aromatic hydrocarbons, in particular, benzene, toluene, ethylbenzene, and xylene (BTEX). This contamination can originate from a variety of sources but is often due to fossil fuel processing and retailing. These toxic pollutants pose a serious health risk to humans, as well as any ecosystems they interact with. As a result, they need to be broken down or removed from the environment. Organisms that can degrade BTEX compounds will be isolated, and their growth in the presence of BTEX compared. Proteomics, functional genomics, and enzyme kinetics will help identify the best degrading enzymes. Intermixed and optimised degradation pathways that can be expressed within a model organism such as E. coli will be designed and tested for use in the bioremediation of BTEX pollution sites. The final goal is to build a biosensor for the detection of BTEX compounds to indicate when they are present and when the site has been sufficiently remediated.
Enmodulators - UTS
Current vaccines for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been proven to be effective against the early Alpha variant but offer lower protection against emerging variants of concern (VOC). Additionally, current vaccine options come with highly specialised manufacturing processes, cold-chain supply issues and limited storage time that make them less accessible to low-resource regions. This has resulted in a lower vaccination rate and more infections in these regions. To tackle this global health challenge, ‘The Enmodulators’ from UTS, aim to develop a cheap, stable, and easy-to-manufacture vaccine platform to rapidly respond to VOCs as they arise. To achieve this, we will repurpose natural protein nanoparticles into a plug ‘n’ play platform that allow the controlled attachment and simultaneous display of multiple SARS-CoV-2 antigens. If successful, this SynBio project could breakthrough existing barriers and provide a new strategy to immunize populations against SARS-CoV-2 in low-resource regions.
Incentivus - QUT
Incentivus, the factory for production of the sesquiterpene aggregation hormone and fragrance. Sesquiterpenes are a class of 15-carbon terpenes that commonly occur in nature, i.e., stink bug aggregation hormones and fragrance molecules from plant essential oils. Incentivus aim to prototype high-level production of these molecules using yeast cell factories. We have cloned 11 sesquiterpene synthases, which separately produce (1S,6S,7R)-sesquipiperitol, (Z)-alpha-bisabolene, (S)-beta-bisabolene, beta-farnesene, alpha-farnesene, cis-muuroladiene, alpha-santalene, and valencene. These sesquiterpene synthases will be introduced into S. cerevisiae with the mevalonate pathway overexpressed. The production of target molecules will be examined through yeast fermentation and instrumental analysis. The aggregation hormones can be used as non-insecticide pest control agents to mitigate infestation of stink bugs. Moreover, 11 sesquipterpene synthases will allow us to investigate the productivities of yeast factories for different products. The data can be integrated into the economy models of biomanufacturing as opposed to chemical production and extraction from plant tissues
Melbourne Macrophages - UMELB
In chronic inflammatory diseases, and as we age, we see an increased ratio of inflammatory to anti-inflammatory macrophages. We are looking to devise a way to transform inflammatory macrophages to anti-inflammatory
macrophages, to reinvigorate tissue repair function and reduce inflammation. We aim to employ genetic intervention on iMACs, to deliver genes involved in anti-inflammatory activation, only expressing them if they are in inflammatory macrophages. This requires the use of a specific biosensor. Thus, the Melbourne Macrophages team have been focussing on designing nucleic acid sensors that can differentially express genes in inflammatory macrophages based on their unique micro mrna profiles, using mi-RNA expressed in high levels and mi-RNA expressed in low levels in inflammatory cells as activators and inhibitors of gene expression, respectively.
Nanobuddies - USYD
Nanobodies are the epitope binding fragments from camelid heavy-chain antibodies. They possess all of the antigen-binding qualities of typical antibodies, but are smaller, more robust, and have lower immunogenicity. These traits result in nanobodies finding implementations across many diagnostic, therapeutic and research applications such as tumour-targeting therapies and ELISA assays. However, there are downsides to nanobodies, including the use of live animals as hosts for production, and the long periods of research time needed to develop new nanobodies to keep up with rapidly mutating or highly diverse antigen targets (e.g. SARS-CoV2). Nanobodies can be functionally expressed in easy-to-grow heterologous hosts such as bacteria, which has huge advantages for the speed of development and manufacturing, in addition to eliminating animal ethics issues. Our project aims to take advantage of this, by using synthetic biology methods to evolve novel nanobodies in vitro via DNA shuffling. This involves recombination of genes in vitro to generate large mutant libraries, which can be screened using E.coli surface display to search for variants with desirable properties, such as increased affinity, specificity or binding to novel epitopes. This process will allow nanobodies to be rapidly reevolved and redeployed to create e.g. updated rapid flow tests (RATs) which can detect new coronavirus variants.
NanoCannoli - UNSW
Biocatalysis is becoming an increasingly important tool in the synthesis and processing of pharmaceutical and chemical compounds, mainly due to its cost-effectiveness, high production yields, eco-friendliness and stereoselectivity. Although many biocatalytic approaches have been developed in the past few years, the efficiency of industrial bio catalysis is nevertheless lower when compared to catalysis in nature. NanoCannoli aims to improve this efficiency by controlling liquid-liquid phase separation (LLPS) in nanoscopic containers. LLPS droplets may attract high local concentrations of substrate and enzymes, thus imitating the way in which biology enhances the efficiency of multi-enzyme reactions. The system involves attaching several multi-valent protein constructs purified from bacteria onto a DNA origami structure and adding cognate proteins and monovalent substrates to form a LLPS system. This would allow for the compartmentalisation of these protein – protein interactions to occur. Following this, single-molecule fluorescence microscopy will be used to examine and validate the LLPS system.
nANUbodies - ANU
Single-domain antibodies, known as nanobodies, are advantageous for use over antibodies in many research, diagnostic and therapeutic fields due to reduced size and increased stability. These stable and soluble alternatives are derived from the single heavy chain binding domain in camelid antibodies. However, current nanobody use has been limited due to individualised production methods for specific functions.
To increase the efficiency and reduce the price of nanobody production we aimed to develop a modular ‘plug and play’ cloning platform to express nanobodies in bacteria and plant expression systems. To this end, in our project we are producing existing nanobodies against well characterised protein tags and proteins of interest. Fluorophore conjugation and fluorescent protein fusions will allow us to evaluate and optimise the function of our nanobodies and the efficiency of our bacteria/plant-compatible production system.
We are developing computational methods to engineer a stable and efficient nanobody ‘chassis’ that can be modified for different functions, which can be used to produce nanobodies against a wide range of targets without animal inoculation. In the future we aim to use our production system to investigate novel nanobody targets for diagnostic or therapeutic applications.
In parallel we have identified conjugation sites on the nanobodies to measure the kinetics of binding reactions in order to identify ideal conjugation reagents. We are introducing mutation sites in nanobodies as a means for conjugating small molecules like fluorescent dyes and drug molecules. In the future we hope to pair these conjugation techniques with the computational development of nanobodies against therapeutic targets resulting in a highly specific drug delivery system.
ResolvSyn - ANU
Resolvins are fatty acid metabolites and are powerful anti-inflammatory and immunoregulatory lipid mediators. They play an important protective role in chronic inflammatory diseases and sepsis - a life-threatening complication of an infection. Producing resolvins requires the enzymatic metabolism of fish oil's two major omega-3 fatty acids; docosahexaenoic and eicosapentaenoic acid (DHA/EPA). Due to the low availability of resolvins from natural sources and inefficient synthesis methods, we have employed synthetic biology to genetically engineer a novel yeast strain that synthesises DHA/EPA into resolvins. This yeast-based biofactory is fuelled by high yielding DHA/EPA algae to sustainably and affordably produce resolvins.
GMBros - QUT
The oleaginous yeast Yarrowia lipolytica has exhibited notable qualities for industrial use due to the natural hydrolysis of triglycerides into fatty acids and glycerol. The aim of QUT synbio team, GMBrOs is to genetically modify this organism for use as a skincare product, able to pass regulatory requirements for a live therapeutic and be released into the environment safely.
An auxotrophic marker was introduced to prevent GMO escape by knocking out the LEU2 gene, removing the ability for Y. lipolytica to produce leucine. The SUC2 gene from S. cerevisiae was knocked into the LEU2 position on the Y. lipolytica genome by homologous recombination. This allowed the secretion of invertase and utilisation of sucrose as a carbon source by the yeast. By applying sucrose for the biotherapeutic to consume, it will be able to outcompete the native skin microbiome.
To test for our strain’s capability for recombinant protein expression, yEGFP was transformed into the pCRISPRyl plasmid Cas9 site by Gibson assembly. Protein expression and SUC2 transformation will be analysed by phenotypic assay and sanger sequencing.
PROTECC Coral - UNSW
PROTECC Coral (Prevent Reactive Oxygen and Thermal Extreme Caused Carking)
The Great Barrier Reef is the world’s largest coral system, integral to Indigenous Australian culture and classified as a World Heritage Site. Rising ocean temperatures have caused several large coral bleaching events, which are attributed to a shift in the symbiotic relationship between coral and microscopic algae species. Heat-induced oxidative stress experienced by algae eventually leads to their expulsion from coral.
PROTECC Coral aims to reduce coral bleaching by increasing the thermotolerance and antioxidant capacity of a common algal symbiont Symbiodinium goreaui. The twofold solution involves introducing small heat shock proteins to prevent protein aggregation, and a glutathione recycling enzyme system to counteract oxidative stress. Experiments and computational modelling were conducted to examine and validate the solution, complemented by considerable outreach, both informing the wider population about synthetic biology and consulting with various stakeholders, including Traditional Owners, to assess the value and impact of the project.
RNA MEChanics - UNSW
Multi-enzyme complexes (MECs) are one of nature's ways of optimising biochemical reactions. We are the RNA MEChanics and aim to custom-design MECs for specific reactions in order to improve their efficiency, yield, and lower cost. We do this by using genetic material (RNA) as a scaffold to bind with high affinity to PUF RNA binding proteins. The modular nature of our model means we can adapt our designs to work with different enzymes from different reaction pathways. We have been testing our system with the biosynthetic pathway for an important redox cofactor F420.
DeNovocastrians - UON
Our project aims to tackle the emerging environmental and health hazard of microplastic pollution. The goal of our system is to degrade microplastics into less harmful compounds and prevent them from cycling through ecosystems. We have designed a functional microbial scaffold that will secrete engineered plastic degrading enzymes. We successfully cloned engineered constructs of the plastic degrading enzymes, MHETase and manganese peroxidase, containing secretion factors and binding domains specific to our scaffold, into Escherichia coli. We envision our system being applied to remove microplastics from aquatic environments such as the ocean, rivers, and water treatment plants.
Phage Phriends - UQ
Urinary tract infections (UTIs) are one of the most common health-care infections, especially in the case of indwelling catheters. In 50-90% of UTIs, E. coli are the most common organism observed. Biofilm formation occurs in approximately 62% of uropathogenic E. coli (UPEC) and contributes to their pathogenicity and antibiotic resistance.
Bacteriophages have evolved to infect specific host bacterial strains and self-replicate whilst producing lytic enzymes such as lysins and depolymerases. These enzymes destroy the protective biofilm matrix. Our project aims to use Synthetic Biology to introduce visually detectable chromoproteins into phage genomes as a biosensor for UTIs. The purpose of this is to reduce the burden of UTI’s by allowing for early detection which would make for more effective treatments.
Free Coli - USYD
Free Coli aimed to improve one of synthetic biology's foundational technologies by designing a genetically modified naturally transformable lab strain of Escherichia coli to produce a more affordable and efficient host organism that does not require chemical treatment or electroporation to become competent. Literature on natural transformation in E. coli and related bacterium was reviewed and experts were consulted to identify twenty-five putative natural transformation genes in A. baylyi for insertion into E. coli via a novel recombineering strategy for insertion of multiple gene clusters. K-means clustering used existing data on transcriptome concentration and promoter strength to model the optimal clustering of genes into eight <5kB DNA fragments. Bioinformatics analysis was conducted to assemble the genes into fragments with salicylate promoters and selectable markers. The wider community was engaged and consulted to affirm the project’s purpose and align its future implementation with the UN’s Sustainable Development Goal to ensure quality education for all.
Design Lab - UTS
Synthetic biology is at the cutting-edge of innovation and technological development, however, most of the wicked problems that society faces are far too complex to be tackled by a single discipline alone. Instead, these challenges require technical skills and expertise from disparate fields of knowledge to be cleverly merged with one another. Our Aus SynBio Challenge team was comprised of scientists and designers, and demonstrated the potential of large-scale interdisciplinary collaboration. Together, we created the electronic magazine (e-zine) “Synthesis” - a curated collection of articles that explores current social issues, scientific and industrial advances, and collaborative co-creation between synthetic biologists and biodesigners. The E-zine reflects our team’s journey throughout the Aus SynBio Challenge, representing the varying challenges we encountered as an interdisciplinary team and the successful solutions we conceived together along the way. “Synthesis” serves as an educational tool and creative output, showcasing the power of interdisciplinary collaboration, and inspiring both the scientific and design communities to immediately seize upon the opportunity to work synergistically on ambitious projects to overcome current and future societal issues.
ProAMP - UWA
Burkholderia pseudomallei (B. pseudomallei) is a prevalent environmental bacterium that causes melioidosis. To combat this disease, the UWA ProAMP team aims to develop a probiotic with the ability to release targeted antimicrobial peptides (AMPs) against B.pseudomallei. Ubonodin, an AMP originating from B. ubonensis with antimicrobial specificity towards Burkholderia strains, will be used as a model for the synthesis of a novel AMP library screening against B. pseudomallei. The introduction of a functional AMP with appropriate biosensing modules into a probiotic species could allow the pathogen to be targeted upon entry into the body, removing the threat of melioidosis without harming the host microbiome.
Plants in Space - UWA
NASA has announced plans to set up a Lunar base in 2028 and a subsequent human crewed mission to Mars in the 2030s. Long term habitation in space requires access to nutritious food in an environment where crops and livestock cannot be grown. The UWA Plants in Space team are engineering moss to produce proteins with increased nutritional value for astronauts on long term space missions. Specifically, we are expressing proteins that are enriched for essential amino acids that could be a supplement for both Earth and Space applications. Moss has the potential to be a sustainable, multifunctional protein biofactory.
We aim to address ocean microplastic pollution through the genetic modification of Vibrio natriegens (V.natriegens), a fast-growing marine bacterium which thrives in saline environments. We have inserted several genes encoding PET plastic-degrading enzymes into V.natriegens via a modular cloning reaction and carefully executed genetic design. In combination with V.natriegen’s natural ability to form biofilms, our engineering methods will provide the modified bacterium with every opportunity to capture and degrade microplastic particles in marine environments, with a precision unmet by conventional bioremediation techniques.
The QUT SynBio team are assessing six novel Australian isolates of Yarrowia lipolytica for their ability to produce pyomelanin. Promising isolates are being transformed for increased pyomelanin production through the knockout of HmgA1. Pyomelanin has multi-industry applications surrounding its photoprotective potential and use in the synthesis of gold nanoparticles.
The SP1 spike protein of SARS-CoV-2 virus binds to ACE2 receptor to gain entry to the host cell. We aim to engineer macrophages to produce soluble ACE2 over an extended period as a possible COVID19 therapy. A modified ACE2 sequence will be introduced to the GAPDH gene locus of induced pluripotent stem cells which will then be differentiated into macrophages that can constitutively express soluble ACE2. We are also investigating the regulation, cost, access, scalability, and public perception of our project through the lens of a similar therapy that has advanced to clinical use: CAR-T cells. Through a series of articles, interviews and social media posts, we hope to communicate the science and challenges of CAR-T cells to the public and reflect on how we can tackle these in our own project to ensure an effective, safe, equitable treatment that meets the needs of the patients it aims to benefit.
Cryptoccocal meningitis is the leading cause of death for HIV positive individuals worldwide, particularly in resource-limited regions in Africa. We are using the latest tools and methods based on CRISP/Cas9 gene editing and GoldenGate cloning to modify bakers yeast (Saccharomyces cerevisiae). We aim to engineer this microbe into a biological sensor for the pathogenic fungus that causes the infection (Cryptococcus neoformas) to create a fast, accurate and cheap diagnostic tool for AIDS patients worldwide.