The Dawn of a New Era in Synthetic Biology at NUS

In a groundbreaking advancement from the National University of Singapore, researchers have developed a revolutionary platform that accelerates the evolution of bacteria to perform intricate tasks, such as degrading plastics. This innovation, known as Lytic Selection and Evolution or LySE, promises to transform how scientists engineer microbes for environmental cleanup, pharmaceutical production, and sustainable manufacturing. Led by Assistant Professor Julius Fredens and PhD candidate Shujian Ong from the NUS Yong Loo Lin School of Medicine's Department of Biochemistry and the Synthetic Biology for Clinical and Technological Innovation programme, or SynCTI, the method bridges the gap between slow, precise genetic engineering and rapid, uncontrolled evolution techniques.

The core challenge in synthetic biology has long been optimizing multi-gene pathways—groups of genes that work together like an assembly line to process complex molecules. Traditional directed evolution, where bacteria are mutated and selected over generations for desired traits, often mutates the entire genome, leading to unwanted side effects or inefficient results. LySE changes this by focusing mutations precisely on target gene clusters while speeding up the process dramatically.

Demystifying Directed Evolution in Microbial Engineering

Directed evolution mimics natural selection in the lab. Scientists introduce random mutations into bacterial DNA, expose the population to selective pressures—like a chemical they must metabolize to survive—and pick the survivors for the next round. For single enzymes, this works well, but for pathways involving multiple genes, like those needed to break down plastic polymers into usable components, it's cumbersome. Pathways can span thousands of DNA base pairs, and evolving them requires balancing expression levels, enzyme activities, and regulatory elements simultaneously.

At NUS, the team recognized that existing continuous evolution systems, such as phage-assisted continuous evolution, or PACE, generate mutations too chaotically and are limited to smaller DNA segments. Discrete methods offer control but are too slow for industrial timelines. LySE combines the best of both: hyper-fast mutation rates controlled at discrete checkpoints.

How the LySE Platform Works: A Step-by-Step Breakdown

The ingenuity of LySE lies in repurposing the T7 bacteriophage, a virus that infects Escherichia coli bacteria. Here's how it unfolds:

• Setup the Phagemid: The target gene cluster—up to 40 kilobases—is inserted into a phagemid, a small DNA ring packaged into phage particles alongside the viral genome.
• Infection and Hypermutation: Engineered T7 phages infect bacteria expressing an error-prone T7 DNA polymerase. This polymerase mutates the phagemid DNA at a staggering rate—160,000 times higher than the bacterium's own replication—introducing diverse changes primarily in transitions (A to G, C to T).
• Lysis and Packaging: The phage lyses the cell in minutes, packaging mutated phagemids into new particles. High multiplicity of infection, or MOI, ensures intense mutagenesis.
• Transduction and Selection: Low MOI phages transduce fresh bacteria. Under selective conditions (e.g., plastic-derived ethylene glycol as sole carbon source with diminishing glucose), improved variants thrive.
• Iteration and Transfer: Repeat cycles select top performers. Optimized clusters transfer cleanly to new hosts without off-target baggage.

This cycle yields precise, potent improvements. In tests, sequencing revealed mutations in promoters boosting expression and enzyme tweaks enhancing activity.

Proof-of-Concept: Bacteria Trained to Devour Plastic Building Blocks

To demonstrate LySE's power, the NUS team targeted a nine-kilobase pathway for metabolizing ethylene glycol, or EG—a key PET plastic component. PET, used in bottles and packaging, pollutes Singapore's waters and landfills. Starting bacteria grew poorly on EG; after five LySE cycles, the best strain produced 50.9 percent more biomass, a 2.8-fold growth boost over controls.

Mutations included a promoter tweak upregulating genes 200-fold and amino acid changes in enzymes like Gox0313 (Y94F) and glcD (I312F), validated individually for contributions. Unlike traditional adaptive laboratory evolution, or ALE, where bacteria cheat by scavenging glucose, LySE enforced true pathway optimization.

A secondary test evolved tigecycline antibiotic resistance via the tetA gene, achieving a 25-fold increase—proof of versatility.

SynCTI at NUS: Pioneering Singapore's Synthetic Biology Landscape

SynCTI, directed by Associate Professor Matthew Chang, positions NUS as Asia's synbio leader. The programme engineers microbes for therapeutics, diagnostics, and sustainability. LySE exemplifies this, building on prior work like bacteria for targeted chemotherapy and gut ammonia clearance. NUS's interdisciplinary ecosystem—spanning medicine, engineering, and computing—fuels such innovations, supported by Singapore's Agency for Science, Technology and Research, or A*STAR.

Singapore universities like NUS and Nanyang Technological University, or NTU, invest heavily in biotech amid national pushes for a green economy. NUS ranks top in Asia for biological sciences, training students in CRISPR, metabolic engineering, and now advanced evolution tools.

Singapore's Plastic Pollution Crisis and the Need for Microbial Solutions

Singapore generates over 900,000 tonnes of plastic waste yearly, with marine debris a growing threat in its busy straits. Microplastics harbour pathogens, impacting fisheries and health. NUS research shows plastics colonised by toxic bacteria like Vibrio, exacerbating coral bleaching.

Current recycling recovers just 20 percent; biodegradation via enzymes like PETase shows promise but needs pathway-scale optimisation. LySE-trained bacteria could upscale plastic-to-fuel or chemical conversion, aligning with Singapore's zero-waste vision by 2030.

Challenges Overcome and Advantages of LySE

• Scale: Handles 40kb clusters vs. PACE's 8kb limit.
• Control: Discrete pauses prevent cheaters; fresh hosts discard junk mutations.
• Speed: Five cycles rival months of ALE.
• Transferability: Optimised phagemids move to industrial strains like Pseudomonas.
• Accessibility: Standard labs suffice—no fancy microfluidics.

Patents filed signal commercial trajectory.

Expert Perspectives and Quotes from the NUS Team

Assistant Professor Julius Fredens notes, “LySE lets us hit pause to control evolution and avoid errors.” Shujian Ong explains the phage's role: “It packs mutated genes for precise inheritance.” Fredens adds on plastics: “Bacteria mutate wildly without focus; LySE hones the pathway cleanly.” Future-wise: “Optimising AI-designed CO2 pathways is massive potential.”

Implications for Biotechnology Careers in Singapore

Synbio jobs boom at NUS, with roles in pathway design, phage engineering, and scale-up. Singapore's biotech sector, valued at SGD 40 billion, seeks PhDs in microbiology and bioinformatics. LySE opens doors to startups tackling plastics, drawing global talent.

Future Horizons: From Lab to Global Impact

The team eyes carbon capture and novel therapeutics. NUS plans LySE workshops, training Asia's next synbio leaders. Amid climate urgency, this positions Singapore universities as biotech powerhouses.

Singapore's Higher Education Edge in Sustainable Biotech

NUS exemplifies Singapore's higher ed focus on real-world solutions. With NTU's synbio labs and SMU's data science, the ecosystem fosters interdisciplinary talent. Government grants like the SGD 1 billion AI fund amplify research.

Students gain hands-on evolution projects, preparing for green jobs. As plastics choke oceans—8 million tonnes enter yearly—LySE-trained microbes offer hope, blending academia and industry.

Photo by Ali Shah Lakhani on Unsplash