ECO-Casein/iGEM NCHU 2025/description

Our project addresses the critical environmental challenges of traditional livestock farming through synthetic biology, creating a sustainable microbial platform for dairy protein production that significantly reduces greenhouse gas emissions while maintaining nutritional value.

The Climate Challenge of Livestock Farming

Global Emission Impact:

FAO Estimate (2023): Livestock contributes 12% of global GHG emissions
Breakthrough Institute (2023): Up to 19.6% depending on methodology
Cattle Dominance: Responsible for 62% of livestock emissions
Primary Sources: Enteric fermentation, manure management, feed production

Current Mitigation Strategies

Energy & Resource Management

Farm energy efficiency improvements
Renewable energy adoption
Soil carbon sequestration

Animal-Oriented Strategies

Selective breeding for efficiency
Feed modification to reduce methane
Improved animal welfare

Systemic Improvements

Precision feeding optimization
Dietary change promotion
Food waste reduction

Waste Utilization

Manure management improvements
Biogas production from waste
Nutrient recycling systems

While existing measures can mitigate emissions to some extent, most remain focused on optimizing existing livestock systems rather than fundamentally transforming the animal-based production model. Our project addresses this limitation by creating a new paradigm for dairy production.

Problem Statement & Project Goals

The Fundamental Challenge

Limitations of Current Approaches:

Focus on emissions reduction per unit rather than system transformation
Inability to address inherent high-carbon nature of livestock production
Dependence on animal-based production models
Limited scalability of incremental improvements

Our Innovative Approach

We explored whether it's possible to bypass animals entirely and use microorganisms to produce essential dairy proteins. Starting with milk as our focus, we aim to create dairy alternatives that balance both nutritional value and environmental sustainability.

Project Objectives

Microbial Protein Production

Use Kluyveromyces marxianus to express functional A2 β-casein, producing digestively friendly milk protein

Carbon Fixation Integration

Introduce carbon fixation enzymes to enable yeast to recycle metabolic CO₂, reducing net carbon emissions

Health Awareness Promotion

Communicate the benefits of A2 β-casein and its potential in low-carbon dairy alternatives

Human Practices Integration

Enhance public understanding and acceptance through outreach and expert engagement

Our Synthetic Biology Solution

The EcoCasein Platform

Core Technology: PGASO System

Platform: Promoter-based Gene Assembly and Simultaneous Overexpression
Advantage: Simultaneous insertion and expression of multiple target genes
Efficiency: More precise than traditional step-by-step transformation
Application: Ideal for complex pathway integration

Dual-Module Design

Protein Production Module

Target Protein: A2 β-casein
Health Benefit: No BCM-7 production during digestion
Gene Sources: Bos taurus (cow) and Bos grunniens (yak)
Host: Kluyveromyces marxianus

Carbon Fixation Module

Function: Convert metabolic CO₂ into usable sugars
Benefit: Carbon emission recycling
Efficiency: Reduced substrate consumption
Sustainability: Lower net carbon footprint

Through this integrated design, we establish a sustainable and low-emission microbial platform for milk protein production, demonstrating the transformative potential of synthetic biology in sustainable food innovation.

Key Experimental Results

Successful Implementation & Findings

Genetic Engineering Success

Plasmid Construction & Verification:

Successful construction of recombinant A2B casein plasmids
Accurate ligation confirmed by Sanger sequencing
Dual RuBisCO cassette maintained in K. marxianus 301
High-purity DNA fragments suitable for transformation

Strain Development

Yeast Transformant Groups:

807 Series: A2B + RuBisCO genes
- 807t1–t8: Bos taurus source
- 807m1–m8: Bos grunniens source

812 Series: A2B only
- 812t1–t8: Bos taurus source
- 812m1–m8: Bos grunniens source

Metabolic Performance

Aerobic Growth

RuBisCO-positive strains showed superior stationary phase stability
Enhanced carbon utilization efficiency
Reduced metabolic burden effects

Anaerobic Metabolism

Modified carbon flux in RuBisCO strains
Slower CO₂ increase after 16 hours
Potential carbon reassimilation activity

While successful gene integration and metabolic engineering were achieved, protein detection requires optimization of extraction methods. The freeze-thaw lysis approach may have caused incomplete cell disruption, highlighting the need for improved protein recovery protocols.

Human Practices & Societal Impact

Connecting Science with Society

Comprehensive Engagement Strategy:

Data Collection: Literature reviews and market analysis
Public Engagement: Educational outreach and demonstrations
Industry Consultation: Expert interviews and business planning
SDG Alignment: Direct contribution to UN Sustainable Development Goals

Educational Impact

Huwei High School Outreach:

Synthetic biology concepts introduced through interactive sessions
Hands-on demonstrations of microbial milk production
66.7% of students expressed willingness to try microbial dairy
Educational materials created for continued use

Expert Integration

Professor Wan-Yu Liu Consultation (NCHU Forestry):

Carbon credit systems and biological carbon sequestration
Potential integration with carbon neutrality goals
Business model development for sustainable dairy
Industry transition strategies

Our Human Practices ensured the project evolved from a purely technical initiative into a socially connected, educational, and industry-aware effort—embodying our vision of transforming "lab-scale innovation into societal sustainability."

Future Directions & Vision

Strategic Development Pathway

Technical Optimization

Protein Extraction Improvement

Test enzymatic digestion methods
Implement mechanical disruption (bead beating)
GFP reporter verification of translation
Western blotting for enhanced detection

Metabolic Characterization

Extended anaerobic cultivation studies
Sugar consumption rate analysis
Carbon fixation efficiency assessment
Stationary phase metabolic profiling

Educational & Commercial Development

Public Outreach Expansion

Continued synthetic biology education
School collaboration programs
Accessible teaching materials
Interactive science activities

Industrial Integration

Scalable microbial platform development
Sustainable dairy brand establishment
Carbon-neutral industry transition
Scientific transparency commitment

Through these continued efforts, we envision EcoCasein as a model that bridges science, education, and industry—pioneering new possibilities for a sustainable and carbon-neutral future in dairy production that addresses both environmental challenges and human health needs.

References

Key Supporting Literature

Food and Agriculture Organization of the United Nations (FAO). (2023). Pathways to Lower-Emission Livestock: Strategies for Climate Change Mitigation and Adaptation. Rome: FAO. Available at: https://www.fao.org/3/cc9871en/cc9871en.pdf
Ritchie, H. & Roser, M. (2023). Livestock Don't Contribute 14.5% of Global Greenhouse Gas Emissions. The Breakthrough Institute. Available at: https://thebreakthrough.org/issues/food-agriculture-environment/livestock-dont-contribute-14-5-of-global-greenhouse-gas-emissions

Project Description