- The team was officially formed. Meetings were held to discuss goals and development directions for 2025.

- Conducted extensive literature review. After discussions, initially selected Antifreeze Proteins (AFPs) as the project focus and began designing related experiments.

- Initiated research on protein design. Initially selected mutation as the engineering strategy.

- Creatively introduced synthetic biology to students, faculty, and the public during the campus open day event ("Crabapple Blossom Festival"), designing related cultural and creative products.

- Introduced the preliminary project concept of Antifreeze Proteins to participants and solicited suggestions to refine the project idea.

- Collected survey questionnaires to gather public feedback on potential application directions.

- After multiple discussions with the PI, the team finalized Fashion & Cosmetics as their track. Team member roles were assigned, and a preliminary experimental plan was drafted.

- After literature review and discussions with the PI, selected 11 naturally occurring Antifreeze Proteins from different organisms as research subjects. Synthesized plasmids containing the AFP gene fragments and attempted to induce AFP expression, optimizing fermentation conditions.

- Learned the Ice Recrystallization Inhibition (IRI) measurement method for characterizing AFPs.

- Researched various AFP design strategies and studied the mechanisms of AFP antifreeze action.

- Completed the design of mutation sites.

- Established the basic framework for this year's human practices activities, integrating excellent traditional Chinese culture, and released a series of promotional articles.

- Conducted a synthetic biology popularization activity for young children during a science open day, introducing our project concept and engaging them through a DNA double helix model building activity.

- Based on feedback from HP interactions with the enterprise Snefe, iterated the project from optimizing AFPs to developing an Antifreeze Chassis System. Considering biocompatibility, planned to conduct cytotoxicity tests on AFPs.

- Through literature review, identified the EBY100 yeast surface display system and initiated work on constructing the Antifreeze Chassis.

- Conducted the first round of characterization tests on natural AFPs and point-mutant proteins.

- Addressed enterprise feedback regarding complex protein purification processes, purification difficulties, and low target protein yield post-purification by shifting focus to constructing an Antifreeze Chassis. Successfully constructed chassis for some natural AFPs using the EBY100 yeast display system.

- Shifted strategy from point mutations to inverse folding.

- Began building the igemTJmodel as part of the protein screening pipeline.

- Engaged in in-depth communication and collaboration with the enterprise Snefe to understand pain points and demands in the current cosmetics and skincare industry, seeking project issues and potential improvements.

- Discussed with Yue Xiaoyan and Pang Xinyang to further refine the project design.

- Communicated and learned from John Cumbers, founder of SynBioBeta.

- Focused intensively on experiments, working in groups to construct a series of non-natural Antifreeze Proteins (including domain-swapped and de novo designs) and their corresponding Antifreeze Chassis systems.

- Designed and constructed 4 domain-swapped fusion proteins based on Dry Lab's domain fusion strategy.

- Successfully constructed the remaining natural AFP chassis and two endogenous expression chassis lacking the lectin protein Aga2.

- Selected MUT-4 for chassis construction due to its superior performance compared to other AFPs.

- Attempted IRI characterization on the Antifreeze Chassis systems alongside protein IRI characterization.

- Lacking prior reports on IRI characterization for cell cultures, the Wet Lab optimized methods and conditions, eventually establishing a protocol using 10% glycerol (in PBS) for resuspending cells, and characterized some Antifreeze Chassis.

- Completed the construction of igemTJmodel.

- Completed the setup of the chatbot.

- Based on further literature review, switched the protein design strategy to de novo design.

- Communicated and learned from the company Bloomage Biotech, understanding their latest R&D progress in biomaterials and antifreeze technology, and exploring collaboration possibilities.

- Discussed antifreeze performance issues with Professor Wu Yi, who suggested introducing yeast's natural stress resistance mechanisms.

- Based on feedback from HP interactions with Bloomage Biotech, the team considered introducing an optimized trehalose metabolic pathway into the Antifreeze System to provide dual antifreeze capability, further enhancing the chassis's performance. Also worked on further optimizing and upgrading the Antifreeze Proteins.

- Obtained 5 key endogenous trehalose synthesis genes (Key Gene 1-5) via PCR from yeast strain. Successfully constructed a trehalose plasmid containing these five target genes.

- Transformed the trehalose metabolic pathway plasmid into the EBY100 yeast. Conducted a series of antifreeze capability characterizations on the EBY100-pRS413-trehalose strain, confirming trehalose's potential to enhance the system's antifreeze performance.

- Discontinued construction of the domain-swapped fusion chassis due to poor antifreeze performance results from the previous characterization round.

- Characterized proteins obtained based on the inverse folding strategy (6A8K-IF, 4NU2-IF, 3WP9-IF) and the de novo synthesis strategy (De Novo).

- Guided by Dry Lab, began replacing the target sequence of AFP11 with smaller helical sequences from other proteins. Commercially synthesized and transformed constructs to create strains expressing a series of proteins, successfully building the NEW11-9, NEW11-10, NEW11-A, NEW11-B series chassis.

- Used Molecular Dynamics simulation results for computational characterization.

- Designed smaller molecular weight short peptides based on enterprise feedback.

- Some team members attended the CCiC conference in Beijing, China. Presented the project to other teams from China. Received valuable suggestions from many iGEM team members and improved the project accordingly.

- Communicated and learned from iGEM Ambassador Patrick.

- Introduced iGEM and synthetic biology to high school students participating in the "Summit Plan" research camp and engaged in friendly discussions with them.

- Completed the construction of the Wiki website and uploaded the team's work results to the Wiki.

- Further optimized the Antifreeze System by introducing both trehalose and Antifreeze Proteins into the EBY100 chassis to construct a Synergistic Antifreeze System. Also constructed Multi-site Specific Rearrangement and Multi-site Specific Inversion systems.

- Based on Dry Lab's inverse folding and de novo design strategies, replaced sequences in AFP11 with artificial non-natural helices using the same method, constructing Antifreeze Chassis AFP11-A and AFP11-B.

- Introduced both trehalose and Antifreeze Proteins into the EBY100 chassis to construct the Synergistic Antifreeze System. Also constructed Multi-site Specific Rearrangement and Multi-site Specific Inversion systems.

- Analyzed Wet Lab characterization results for AFPs and Dry Lab computational characterization results. Proposed hypotheses regarding the antifreeze mechanism of AFPs and factors contributing to enhanced antifreeze performance.

- Completed the setup of the model for generating Antifreeze Short Peptides.

- Innovatively attempted engineering and commercialization exploration for the project, progressing from single product formulation to communicating with companies and writing corporate white papers, driving the project forward along the axis of product development and production line design.

- Discussed with Professor Mi Wei from the Law School regarding current legal regulations and technical barriers for the application of Antifreeze Proteins in cosmetics.

- Discussed with staff from the government heating department the potential application of Antifreeze Proteins in preventing freezing in heating pipelines.

- Attended the National Ice Cream Industry Exhibition at the Tianjin Meijiang Convention and Exhibition Center to gain insights into the commercialization logic of bio-manufactured products.