Contribution

1. Improved the sucrose synthesis efficiency of cyanobacteria, providing a reference for future teams using cyanobacteria as a carbon source supplier.

• In addition to considering the introduction of cscB to enable the export of sucrose from cyanobacteria, we also considered the overexpression of the sucrose synthase gene sps and the related gene glgC from PCC 7942 to significantly enhance the secretion of sucrose by cyanobacteria. Our results can provide a reference for future teams that wish to experiment with co-cultivation of cyanobacteria and any other organisms.

2. Tested the effectiveness of cultivating cyanobacteria without specialized equipment, encouraging more future iGEM teams to choose it as a chassis organism.

• During the human practice phase, we learned that some iGEM teams did not choose cyanobacteria, a very promising chassis organism, due to the lack of essential lighting incubators or unfamiliarity with related operations. To address their concerns, we explored the effectiveness of cultivating cyanobacteria without specialized equipment and found that this is not an issue. Our results can encourage more iGEM teams to experiment with cyanobacteria.

3. Introducing Wisconsin Fast Plant, a rapid-cycling relative of Arabidopsis, as a superior complementary model plant.

• In plant biology research, while model plants like tobacco and Arabidopsis have been specifically designed to facilitate experimental studies, their cultivation remains both time-consuming and technically demanding. These plants often require precise growth conditions, extended life cycles, and careful maintenance, which can slow down the pace of research. Moreover, the expensive equipment and specialized facilities needed for plant-related projects, such as controlled growth chambers and advanced imaging systems, pose significant barriers to entry, especially for smaller labs or research groups with limited resources. These challenges highlight the need for more accessible and efficient model systems to accelerate discoveries in plant science.

Therefore, we would like to introduce the potential of using Wisconsin Fast Plants, a rapid-cycling relative of Arabidopsis developed by Professor Emeritus Paul H. Williams at the University of Wisconsin-Madison. These plants offer a much simpler operation and boast a significantly faster lifecycle of just 35 days, compared to traditional model plants like tobacco and Arabidopsis.

Additionally, Wisconsin Fast Plants are fully compatible with our TTTT system and exhibit superior performance in leaf spray transfection, thanks to their smooth leaf surface, which lacks the villi or waxy layer that can hinder efficient transfection in other plants. This makes them an ideal alternative for accelerating research and experimentation in plant biology. We are also willing to share the seeds and protocols to future teams.

More information at:Plant

4. For the first time, introduced the HapAmp system to the iGEM community, providing tools for future iGEM teams working with yeast.

• In the engineering modification of yeast, a key question has always been how to efficiently and conveniently increase the yield of target products. This year, we introduced the HapAmp system to the iGEM community for the first time. Using the HapAmp system, we can achieve the simultaneous integration of a metabolite gene cluster with adjustable copy numbers into the yeast genome. This will significantly simplify the time and labor costs for future iGEM teams.

5. For the first time, proposed a suicide switch that balances strain leakage and strain functionality, providing a reference for future iGEM teams focused on environmental-related projects.

• Addressing environmental issues is an important goal within the field of synthetic biology, and many iGEM teams have done significant work in this area. Among these topics, engineering strains to withstand unfavorable conditions in the environment is an unavoidable subject. However, if these engineered strains were to escape, they could potentially harm the natural environment.
  
  Therefore, how to ensure that bacteria can be killed in the event of leakage while remaining stable and controllable for reproduction and functionality in specific environments is a crucial issue that must be addressed. Here, we achieved this important goal by integrating the use of Cre recombinase, the killred gene, and a quorum sensing system.

6. A suicide switch compatible with food environments.

• Food production is also an important topic in synthetic biology, and the design of kill switches needs to be considered comprehensively. We cannot use substances such as iron ions or tetracycline, which are not approved for food use, to regulate the chassis cells in operation. Here, we integrated the use of VanR, a transcription factor regulated by the FDA-approved chemical vanillic acid, along with a NOT GATE to achieve strain leakage suicide while ensuring food compatibility. This approach can be applied not only to different yeasts but also to bacteria.

7. The TTTT system now includes permanent genomic insertion capabilities, expanding its range of applications and goes into proposal of imperal-college iGEM team.

• Conventional approaches for transforming plants encompass the use of Agrobacterium tumefaciens infection, particle bombardment, and viral infection. These techniques have been extensively applied in various model plants, such as Arabidopsis thaliana and tobacco, as well as annual or biennial herbs. In recent years, with the swift progress of nanoscience and nanotechnology during the latter part of the 20th century, nanomaterials have gained widespread adoption in nanobiology and gene therapy.
  
  In the past four years, our team members with Jianhuang's lab at Soochow university focused on the research of PEI-modified CDs (CDP). A type of carbon quantum dot with a positive charge can be implemented through transfection. In iGEM 2023, our team built a project based on it and gave the system a name- Carbon nanodot-based tracked, transformation, translation, and trans-regulation(TTTT) system.

This year, by enhancing the preparation process of CDP, the latest version, known as Smart-CD (patent pending), can effectively transfect plants through leaf spray for transient expression and seed soaking for genomic insertion. This method has been tested on several common crops.

Moreover, we are delighted to see that the imperial-college team has put the TTTT system in their project design and requested material sharing from us. Although they are unable to finish the experiments in this part and unable to present the result this year. We are planning to update this part next year and also looking forward for more collaborations in 2025.

More information at: Plant