Individual Final Project

Carbon Forge Red: Engineering a Photoautotrophic System for the Conversion of CO₂ into L-Lactic Acid as a Raw Material for Poly Lactic Acid on Mars.

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HTGAA 2026: Individual Final Project Documentation

SECTION 1: ABSTRACT

  1. Abstract: Sustainable Mars settlement requires In-Situ Resource Utilization (ISRU) to reduce dependence on Earth-based supply chains. This project addresses the critical need for manufacturing materials on Mars by engineering a biological system to convert atmospheric $CO_2$ into Polylactic Acid (PLA), a versatile bioplastic for 3D printing. The broad objective is to create a photoautotrophic platform using Chlorella vulgaris for carbon fixation and polymer precursor production. We hypothesize that by redirecting metabolic flux from pyruvate to lactate via the introduction of $L$-lactate dehydrogenase ($Lldh$) and pyruvate kinase ($pk$), while knocking down phosphoenolpyruvate carboxylase ($ppc$), significant yields of $L$-lactic acid can be achieved. Specific aims include genetically modifying the algae, validating lactate accumulation, and refining extraction protocols. Methods involve CRISPR-based metabolic engineering, cell lysis, and chromatography for purification, followed by chemical polymerization. This system bridges the gap in Martian ISRU by providing a renewable source for construction and tool fabrication.

SECTION 2: PROJECT AIMS

Define three aims for your final project (minimum one sentence per aim).

  1. Aim 1: Experimental Aim
    The first aim of my final project is to engineer Chlorella vulgaris to produce $L$-lactic acid from $CO_2$ by utilizing CRISPR-Cas9 gene editing to introduce $L$-lactate dehydrogenase ($Lldh$) and pyruvate kinase ($pk$) genes while knocking down the phosphoenolpyruvate carboxylase ($ppc$) gene. I will use Benchling for genetic circuit design and codon optimization, Addgene plasmids for the CRISPR backbone, and Asimov Kernel for metabolic modeling. The experimental workflow involves algal transformation, selection via antibiotic resistance, and verification of lactate secretion using high-performance liquid chromatography (HPLC).

  2. Aim 2: Development Aim
    The second aim is to scale the biological production into a functional manufacturing pipeline by optimizing the downstream purification and polymerization of $L$-lactic acid into 3D-printable Poly Lactic Acid (PLA) filament. Following a successful Aim 1, this phase involves developing efficient cell lysis protocols, utilizing ion-exchange chromatography for high-purity lactic acid recovery, and performing ring-opening polymerization to create a resin suitable for extrusion into 3D printing filaments.

  3. Aim 3: Visionary Aim
    The long-term vision for this project is to establish a self-sustaining In-Situ Resource Utilization (ISRU) framework for Mars settlement, where atmospheric carbon is converted into essential structural materials without Earth-based feedstock. By validating these experiments under simulated Martian atmospheric conditions, this project aims to address the major barrier of high-mass transport costs in space exploration, enabling a new paradigm of “biological manufacturing” where settlers can grow their own tools, spare parts, and habitats from the air they cannot breathe.

SECTION 3: BACKGROUND

Background and Literature Context

Provide background research that explains the current state of knowledge and identifies the gap your project addresses.

  1. Briefly summarize two peer-reviewed research citations relevant to your research (minimum four sentences).
  2. Explain how your project is novel or innovative (minimum three sentences).
    Examples:
    • New applications or uses of existing biological tools or concepts
    • Development of new approaches, methodologies, or technologies
    • Ways the project challenges existing paradigms or assumptions
    • How the work expands the boundaries of synthetic biology
  3. Explain why your project matters and what impact it could have (minimum five sentences).
    Examples:
    • The problem addressed
    • Importance of the problem
    • Broader societal contribution
    • Advancement of knowledge or capability
    • Field-level change
  4. Describe the ethical implications associated with your project and identify relevant ethical principles (minimum two paragraphs).
    • Paragraph 1: What ethical implications are involved?
    • Paragraph 2: What measures should be taken to ensure ethical conduct and societal responsibility?

SECTION 4: EXPERIMENTAL DESIGN, TECHNIQUES, TOOLS, AND TECHNOLOGY

Create a detailed experimental plan for your final project. Include a timeline for each part (minimum 15 lines/sentences).

  • Include specific methods, tools, and technologies for each part of the project and analysis.
  • Describe expected results for each experiment.
  • Include figures if possible to show workflows.
  • Reminder: All HTGAA projects must include some DNA design!

Techniques Checklist

☐ Pipetting
☐ Lab Safety
☐ Bioproduction
☐ Registry of Standard Biological Parts
☐ Chassis Selection (e.g., DH5alpha)
☒ Bioethical Considerations
☐ Plasmid Preparation
☐ Bacterial Culturing
☐ Quality Control/Analysis
☐ Bacterial Processing (Centrifugation, Lysis, DNA Purification)
☐ DNA Construct Design
☐ Restriction Enzyme Digestion
☐ Gel Electrophoresis
☐ DNA Purification From Gel
☐ Cell-Free Systems
☐ Freeze-Dried Cell-Free Systems
☐ Databases (GenBank, NCBI, Ensembl, UCSC Genome Browser)
☐ miniPCR Tools
☐ Protein Purification

Lab Automation

☐ Creating Code for Laboratory Automation
☐ Using Liquid Handling Robots (e.g., Opentrons)
☐ Designing a Twist Order
☐ Creating a plan to use the Autonomous Lab at Ginkgo Bioworks

CRISPR

☐ CRISPR/Cas9
☐ Designing Prime Editing gRNA

Protein Design

☐ Protein Design
☐ Use of Boltz or PepMLM
☐ Use of Asimov Kernel
☐ Use of Benchling
☐ Models and Notebooks
☐ Databases

  1. Expand upon two techniques you checked above (minimum four sentences).
  2. Identify any HTGAA Industry Council companies associated with your project (optional):
    • Addgene
    • Epibone
    • Ginkgo Bioworks
    • Helix Nano
    • Millipore Sigma
    • BioFabricate
    • Biome Consortia
    • Bolt
    • Boltz.bio
    • Cultivarium
    • DeepCure
    • Mycoworks
    • New England Biolabs
    • Opentrons
    • SecureDNA
    • Takeda Pharmaceuticals
    • Thermo Fisher Scientific
    • Transfyr.ai
    • Twist Biosciences
    • Upside Foods
    • Waters Corporation

SECTION 5: RESULTS & QUANTITATIVE EXPECTATIONS

You are required to validate at least one aspect of your final project aims.

Acceptable validations include:

  • Designing DNA relevant to your project
  • Performing PCR or Gibson assembly
  • Creating and performing a cell-free assay
  • Running code or computational analysis
  • Designing and testing DNA constructs via Twist
  1. What aspect of your project did you choose to validate?
  2. Write a detailed protocol of how you validated it.
  3. What synthetic biology techniques did you use?
  4. Present data and analysis (experimental or simulated).
  5. Describe challenges, limitations, and alternative strategies.

SECTION 6: ADDITIONAL INFORMATION

  • List all references cited (bullet list).
  • Create a supply list and budget for your project (bullet list).
    • Supplies, equipment, and budget needed.