Week 1 HW: Principles and Practices

¹

Fungal Biorefinery for Sargassum and Henequen Valorization into Antimicrobials

On the Yucatan Peninsula, various problems associated with coastal pollution have been reported due to sargassum wash-ups that affect fishing production and local tourism. Likewise, henequen-producing areas generate a considerable amount of waste that could be exploited through biotechnological processes ².

This integrated biorefinery approach aims to address waste-related environmental crises, reduce dependence on imported pharmaceuticals, and create a circular bioeconomy model in the Yucatan Peninsula.

Governance/Policy Objectives ³

Objective 1: Ensure equitable distribution of benefits and local capacity building.

Prevent biopiracy and ensure that economic and knowledge benefits remain within local Mayan communities.

a) Legal Frameworks for Resource Sovereignty: Establish clear agreements that define sargassum as a “biological resource” under state sovereignty before recognized institutions (SEMARNAT), with protocols for access and benefit sharing aligned with the Nagoya Protocol.

b) Technology Transfer & Workforce Development: Create mandatory partnerships between national and international biotech companies, as well as universities and federal research centers (UADY, CECIHTI, SIIDETEY), including joint ownership of intellectual property and training programs in genetic engineering and bioreactor operation.

c) Community-Led Monitoring Committee: Form a permanent oversight committee composed of representatives from coastal communities, henequen ejidos, and bioethicists (CONBIOÉTICA) to review project direction, benefit sharing, and environmental impact.

Objective 2: Implement responsible governance regarding environmental release and containment.

Mitigate ecological risks from genetically modified fungi and ensure that the project improves local ecosystems rather than disrupting them.

a) Strict physical and biological containment standards: Require the exclusive use of genetically modified strains with multiple self-destruct switches. The aim is to have genetic safety measures such as genetic microfactories in place to prevent environmental damage.

b) Environmental monitoring based on the precautionary principle: Establish an environmental DNA monitoring network before and after implementation to track any possible leaks and impacts on the native ecosystem.

c) Transparent communication of risks and benefits: Develop risk assessment control panels aimed at the public, departments dedicated to constant updating, and detailing the effectiveness of the strategies developed.

Potential governance actions

1. Establish a Yucatán Bioeconomy Regulatory Sandbox

Regulations on GMOs in Mexico are restrictive and not adapted to genetically modified environmental applications.

That is why I would like to propose the creation of a temporary, legally recognized “Biological Containment Unit” in designated areas (e.g., the Progreso Industrial Park or the Yucatan Science and Technology Park [PCTY]) where suitable facilities allow for pilot-scale testing of genetically modified fungi under close supervision. It should be led by the Ministry of the Environment (SEMARNAT) with CONABIO, CIBIOGEM, and the Yucatán government. The necessary investment can be provided by interested public institutions (e.g., the Yucatan Ministry of Science, Humanities, Technology, and Innovation [Secihti], the Scientific Research Center of Yucatán [CICY], CINVESTAV) in collaboration with biotechnology venture capital firms (e.g., GridX, Zentynel Frontier Investments) for the installation and maintenance of activities.

• Regulators are expected to be willing to adopt adaptive governance models, with 100% effective containment protocols at the pilot scale and positive public opinion managed through transparency.

• The aim is to prevent possible regulatory paralysis or suspension of the testing space before it begins due to mistrust in the public reaction.

2. Launch a “Community Biorefinery” Co-Ownership Model

Waste recovery projects often extract resources without returning profits or control to local collectors/workers.

This proposal is to develop a cooperative ownership structure in which companies specializing in sargassum collection and henequen producers hold shares in the establishment of the biorefinery, participating in its management alongside cooperatives, the Agency for the Development of Yucatán (ADY), private investors, and environmental engineers from the area. The initial capital will come from impact funds, supplemented by support from the Yucatán Ministry of Sustainable Development (SDS), with cooperative assemblies overseeing important decisions.

• It is expected that the cooperatives will have the organizational capacity to participate, that social trust will be generated, and that the return value of potential products will be compelling enough to generate meaningful participation.

• There is a risk that in the future the cooperatives will be marginalized in technical decisions, which would lead to conflicts and disruption of the supply chain. Another scenario suggests that commercialization could lead to overexploitation of sargassum or the diversion of henequen from traditional artisans, causing cultural erosion.

3. Create an Open-Source Fungal Toolkit for Safe Engineering

Advanced synthetic biology tools (e.g., CRISPR vectors, biocontainment modules) are patented and expensive.

Given the lack of organization of genetic data on fungi in the region, I would like to propose the establishment and development of an open-source repository of standardized genetic components (BioBricks) through a recognition and optimization program for filamentous fungi, freely accessible to all partner institutions and the interested scientific community. The initiative will be carried out in collaboration between public universities (CINVESTAV, UADY, TECNM, UNAM) and private universities (Anáhuac Mayab, Marist University, Modelo University), with technical contributions from international synthetic biology institutes (e.g., Ginkgo Bioworks), funded by Secihti and philanthropic organizations (corporate foundations such as Carlos Slim, Harp Helú, and Gonzalo Río Arronte). Access will be public through a locally hosted digital platform with material transfer agreements that require bioinformatics security and benefit-sharing principles.

• It is expected that the necessary open-source tools will be available to reduce barriers for local actors, as well as the implementation of infrastructure to maintain databases; compliance with intellectual property standards for partners, communities, and institutions will be monitored responsibly.

• There is concern about the lack of resources and technical protocols for this proposal, resulting in a functionally inferior tool or one with vulnerabilities to the data entrusted to it. The worst-case scenario is that the platform will not be accessible, or that it will lack oversight tools, allowing for the design of risky or unethical experiments.

4. Implement an Independent, Transparent Bio-Impact and Benefit Audit System

Impact assessments are conducted privately, on a one-off basis, by experts hired by the companies themselves. Often, the results are not shared in full with the public or are manipulated.

I would like to suggest the creation of a permanent, independent group responsible for conducting ongoing public oversight of the actual safety of biotechnology projects within the state, in order to provide public transparency regarding the actions and resources used in this and future projects, as well as the benefits to the community. The office responsible for auditing should include partners from the fields of science, technology, and the environment, experts in social issues, and community representatives, with support from national and international organizations. The section should have certifications, a simple public website for accessing reports, and mixed financing between public grants and income generated by specialized services for the rest of the associations.

• The aim is for the company to be transparent with auditors, providing complete data and full access to facilities. Similarly, there is public interest in the publication of progress and the consistency of actions to be taken with the objectives set.

• There is a risk that audit reports will be completed but then dismissed by politicians or businesspeople; that the tests will not detect a risk or social harm, creating a false sense of security; that the audit process becomes so strict and slow that it paralyzes the project’s ability to adapt and improve, stifling innovation; or that the data is used by opponents to create misleading scandals and unfairly damage the project’s reputation.

What is the best proposal?

Taking into account the facilities, the material, economic, and legal resources, as well as the experience gained in other experimental lines related to the handling of genetically modified organisms, I propose to begin with the creation of the “Biological Containment Unit” as part of the start of the economic regulatory sandbox aimed at providing a safe space where, in the future, the design and implementation of experiments and pilot tests with genetically modified fungi can be planned.

In turn, I would seek to complement the technological facilities in access to genomic databases and “biobricks” through agreements with solid and recognized institutions (for example, the European GMO Initiative for a Unified Database System [EUginius], the iGEM Foundation, The National Center for Biotechnology Information [NCBI]) for the collection, experimental management, and storage of genomic data necessary to process in silico trials that support the transition to in vitro experiments.

The proposal is intended for review by Secihti, the Directorate of Innovation and Smart Government of the City Council of Mérida, Yucatán, the Global Biotech Revolution (GBR), the iGEM Foundation, the International Gene Synthesis Consortium (IBBIS), and any other organization interested in biotechnological advances involving filamentous fungi.

Remarks

• During this exercise, I continue to wonder how modifying more complex fungi will really impact industry and daily life. I am concerned that the appropriate management of species and genetic data will not be achieved in the future, which in popular culture could refer to failures such as in the video game “The Last of Us.”

• I have reflected that these types of projects can be a watershed in the safe management of local resources, as long as there is trust with ethnic groups, productive groups, associations, business groups, and non-profit organizations. I would like the technological transition, bioethical, and legal frameworks to develop strategies to update and adapt to possible issues that may arise during the design of projects in this category.

Week 1 HW: Assignment (Week 2 Lecture Prep)

Homework Questions from Professor Jacobson:

1. Nature’s machinery for copying DNA is called polymerase. What is the error rate of polymerase? How does this compare to the length of the human genome. How does biology deal with that discrepancy?

Human DNA polymerase has a common error rate of one error per 10⁶ bp inserted. If the human genome has a length of ~3.2 x 10⁹ bp, approximately 3,200 errors are expected within the entire genome. This does not take into account that the cell has polymerases capable of correcting these errors during replication through a proofreading system, followed by mismatch repair (MMR), base excision repair (BER), and nucleotide excision repair (NER).

2. How many different ways are there to code (DNA nucleotide code) for an average human protein? In practice what are some of the reasons that all of these different codes don’t work to code for the protein of interest?

Given that a protein generated by the human body has an average of 1036 base pairs, when organized by codons, it can be translated into a protein of 345 amino acids; remembering that the genetic code is degenerate (the same amino acid can be encoded by more than one codon) and that there are 61 known codons that encode an amino acid (not counting 3 stop codons), and the complexion for the total number of different ways to arrange N codons of Q different aminoacids is given by:

$ F(N, Q) \approx N \ln(N) - Q^* [(N/Q) \ln(N/Q) - N/Q] $

When we want a code to express a protein of interest, not all of the above combinations will work, as they depend on processes during translation such as the stability of the messenger RNA strand, whether it forms secondary structures, the presence of signals or regulatory elements; the availability of ribosomes and tRNA, as well as the speed of recognition of the start codon and the Kozak sequence (for eukaryotic cells).

Homework Questions from Dr. LeProust:

1. What’s the most commonly used method for oligo synthesis currently?

According to the presentation, I understand that the standard method is solid-phase oligonucleotide synthesis using phosphoramidites in a four-step sequence: phosphoramidite coupling, capping of unreacted sites, oxidation, and deblocking. However, specialized equipment has now been developed, such as the High-throughput oligo synthesizer (Oligator) by Illumina, Inkjet-based DNA microarray by Agilent and Electrochemical-based microarray by CombiMatrix.

2. Why is it difficult to make oligos longer than 200nt via direct synthesis?

The main difficulty lies in the imperfect coupling efficiency in each cycle. Efficiency per step is typically over 99%, however, losses accumulate exponentially with length as each cycle introduces impurities and the stability of the support degrades with prolonged cycles.

3. Why can’t you make a 2000bp gene via direct oligo synthesis?

A linear gene of 2000 bp cannot be manufactured directly and continuously due to the possibility of pairing errors or mutations in the sequence. Furthermore, even if it could be done in a single step, it would be a very costly process. Instead, assembly in Precision Oligo Pools is proposed to improve the signal-to-noise ratio, reduce the burden of oversampling, and increase the representation of the full length.

Homework Questions from George Church:

1. What are the 10 essential amino acids in all animals and how does this affect your view of the “Lysine Contingency”?

There are nine essential amino acids (those that cannot be synthesized de novo and must be obtained from the diet), which are:

• Histidine (H), isoleucine (I), leucine (L), lysine (K), methionine (M), phenylalanine (F), threonine (T), tryptophan (W), and valine (V) ⁴.

The contingency of lysine analyzed from the perspective of metabolism is only a means of regulating the growth of an organism that depends on this amino acid for survival. With synthetic biology, in theory, this limitation can be overcome by implementing genes that encode the enzymes of the metabolic pathway for synthesis, thereby implementing a strategy to overcome this need, which could also be replicated for the rest of the essential amino acids.