In this year's iGEM project, the SR-Shenzhen (ChronoCure) team contributed across multiple levels: new parts, validated data, and conceptual application resources. We submitted 17 parts to the iGEM Registry, including 5 newly designed parts experimentally validated for their roles in NMN and GSH biosynthesis, demonstrating strong feasibility and application potential. We also provided new experimental evidence for the existing part PelB-GLP-1 (BBa_K5083001), enhancing its reliability within the Registry and offering reference data for future peptide secretion studies. Beyond laboratory work, we designed conceptual application resources, including a draft product formulation, a process pipeline, and a biosafety framework, serving as templates for teams interested in probiotics, nutraceuticals, and industrial translation. Finally, as a small high school team, we developed and published a Small Team Management Guideline, establishing a transferable model of efficient collaboration under resource constraints, which may benefit future iGEM high school teams.

This year, we submitted a total of 17 parts to the iGEM Registry, including 5 newly designed parts (Table 1) that were experimentally validated for their feasibility in NMN and GSH biosynthesis. These new parts cover the complete pathway of substrate transport, precursor supply, product synthesis, and secretion, demonstrating both stability and strong application potential. The remaining parts, although derived from these five core modules, also hold scientific validity and practical value. In the future, iGEM teams working on NAD+ metabolism, antioxidant research, or functional probiotic design can directly adopt our parts as reliable tools and foundational elements.

Table 1. Five newly designed parts were experimentally validated by our team.

In this year's project, we conducted new experimental validation for the part PelB-GLP-1 (BBa_K5083001), which was originally submitted by the iGEM24_Squirrel-CHN team for exploring GLP-1 secretion. Based on their design, we reconstructed a plasmid containing the PelB signal peptide fused with the GLP-1 coding sequence (Figure 1A). The PelB-GLP-1 expression cassette is driven by the T7 promoter, with RBS (BBa_B0034) at the 5′ end and a T7 terminator (BBa_B0015) at the 3′ end, forming a complete expression module (Figure 1B). After plasmid assembly, PCR amplification followed by agarose gel electrophoresis revealed a clear band of approximately 162 bp (Figure 1C), confirming the correct insertion and validating the functionality of this expression framework.

Figure 1. Construction and validation of the PelB-GLP-1 expression cassette.

After completing plasmid construction and PCR verification, we further performed a Western Blot (WB) assay to detect the protein expression of PelB-GLP-1. The results showed a specific band below 15 kDa in the target lane (PelB-GLP-1), consistent with the expected size. This indicates that the expression cassette was successfully transcribed and translated in E. coli, confirming the feasibility of PelB signal peptide–mediated GLP-1 expression. Combined with the plasmid construction and PCR results, we provide new experimental evidence for BBa_K5083001, which enhances its reliability in the iGEM Registry and offers valuable reference data for future teams working on peptide secretion systems or GLP-1–related applications.

Figure 2. Western Blot validation of PelB-GLP-1 expression.

Beyond laboratory validation, we also provided practical application resources. We designed a conceptual product draft formulation based on NMN, GSH, and GLP-1, covering both oral supplements and skincare (Figure 3). In addition, we outlined a pipeline from fermentation → purification → biosafety treatment → packaging, and proposed a closed-loop biosafety and compliance framework. Importantly, these resources are presented purely as conceptual designs, with no actual ingredients, product development, or human use involved. They serve solely as reference templates for future iGEM teams interested in exploring functional probiotics, nutraceuticals, and industrial translation pathways.

Figure 3. Conceptual Product Development Draft (Not for Real Use).

As a small high school team, ChronoCure faced multiple challenges during the competition, including limited manpower, heavy workloads, and tight schedules. Unlike many larger university teams, we needed to handle wet lab design and validation, modeling and analysis, Wiki writing and design, as well as public engagement and education—all with a small group of members. To prevent inefficiency caused by poor communication or fragmented responsibilities, we created the Small Team Management Guideline as a unified framework for all members to follow. This guideline not only defined clear roles and responsibilities but also established cross-team collaboration mechanisms, supported by time management practices and a rotation system to ensure both efficiency and flexibility. Most importantly, we documented our experiences throughout the process, creating a transferable resource that offers practical reference for future high school iGEM teams operating under similar small-scale and high-pressure conditions.

We have compiled a “Stakeholder Interview Handbook” aimed at providing future iGEM teams with a standardized process and methodology that can be directly adopted. This manual not only summarizes our own research practices, but also integrates ethical principles and team reflections, helping subsequent teams conduct Human Practices in a more efficient, systematic, and responsible way.

Through this manual, future teams will be able to:

• Get started quickly: Understand the key steps and considerations before, during, and after research;
• Avoid common pitfalls: Reduce errors caused by leading questions, incomplete records, or biased information;
• Ensure ethical compliance: Emphasize informed consent, privacy protection, and transparent traceability;
• Generate actionable feedback: Transform scattered interview content into structured conclusions that provide valuable references for experimental design, product development, and societal outreach;
• Build trust: View research not as one-time data collection, but as the beginning of long-term dialogue and collaboration.

This manual is not only a summary of our team’s work, but also a shared resource and methodological contribution to future iGEM teams.