Week 1 HW: Principles and Practices

Biological Engineering tools:

Rapid Urinary Heavy Metals Ion detection - Paper Based Lateral Flow Assay

Heavy metal ions pose a serious threat to both the environment and human health due to their high toxicity and tendency to bioaccumulate in biological systems. Although conventional analytical techniques such as chromatography offer high sensitivity and accuracy, they are often expensive, and impractical for rapid, on-site detection. Therefore, I plan to develop a portable urinary test the detection of heavy metal ions using modified bioreceptors, such as aptamers, DNA probes, or DNAzymes, to enable selective and sensitive recognition. This approach is inspired by multi-parameter drug test assays, which can simultaneously detect multiple analytes (e.g., amphetamines, morphine, THC, benzodiazepines, and cocaine), using Lateral Flow Assay (LFA) principles.

Bioreceptor design (DNAzyme):

DNAzyme is a DNA molecule that has a specific metal ion-dependent enzymatic activity — that is, it will only perform a catalytic reaction (e.g., cleave a DNA substrate) when the target metal ion is present. DNAzymes have been selected through in vitro selection for their high specificity toward ions such as Pb²⁺, Cu²⁺, and Hg²⁺.

Design outline:

1. Selecting a suitable DNAzyme: Chose a DNAzyme that is specific to the target ion (e.g., Pb²⁺ 8–17 DNAzyme, Cu²⁺, Hg²⁺). The DNAzyme is selected by in vitro selection to achieve high affinity and specificity for a particular ion.
2. Design of cleavable DNA substrates: DNAzymes work by cleaving site-specific DNA/RNA substrates when the target ion is present. The substrates can be visually labeled so the resulting cleavage changes become measurable signals -> AuNP or fluorofor
3. Chemical Modification for integration with biocencor: Add functional groups (e.g., thiol, biotin) to the DNAzyme or substrate to allow it to bind to nanoparticles (AuNPs) or the strip surface. This allows integration with lateral flow system.

Governance/policy goals

Biosensing DNAzyme-based detection intersects with health monitoring, environmental justice, and personal data. Therefore, here is governance goals

1. Responsible & Safe Innovation
   • Transparent Safety validation: clear evaluation of safety, performance, and limitation before deployment
   • Adaptive Regulatory: Following latest monitoring and safety standard
2. Promote Trust, Transparency and Public accountability
   • Equitable distribution: Accessible to diverse populations without socioeconomics barriers
   • Monitor bias and performance: Regulary asssess test performance in different demographic groups, ensuring accuracy and precision
3. Ensure Privacy and Ethical use in implementation
   • Clear informed consent: Ensuring informed consent and secure data storage
   • Limit data use to purpose only: ensure health or exposure data collected by the device is used only for its intended purposes

Potential governance “actions”

Note: Purpose (P), Design (D), Assumptions (A), Risks (R).

1. Establish a risk-based approval path for Rapid Biosensors

   • P: Rapid biosensors often lack clear regulatory pathways, especially for diagnostics like DNAzyme metal sensor. Need to establish a specific risk-based regulation framework for novel biosensors, ensuring performance and validation data are required to clinical use
   • D: Select categories based on use cases (high, medium, low risk) through environmental monitoring. Establish validation standards for complex matrices such as urine. Ensure a review process.
   • A: All regulators have capacity and expertise to evaluate DNAzyme sensors and standarized set of perfomance metrics can be agreed across jurisdictions.
   • R: Failure: Overly stringent regulations, slow transition from research to implementation. If regulations are too strict, product performance quality declines.

2. Funding & Standard for Ethical Design

   • P: Introduce policy incentives and funding programs that require ethical, inclusive design and ongoing evaluation
   • D: Attach ethical & societal impact requirements to grants. Fund interdisciplinary teams (biologists, ethicists, community reps) working together. Develop standards for equitable access
   • A: Funding agencies and researchers value equity and community engagement.
   • R: Increasing project costs/time, disadvantaging smaller labs or startups that lack resources to meet expanded requirements.

3. Public engagement & Transparent report for community or stake holder

   • P: Formalize public and stakeholder engagement requirements during early design, deployment planning, and ongoing use
   • D: Host public consultations, workshops, and panels to integrate values and concern. Open reporting of performance, limitations, and real-world data in accessible formats.
   • A: Stakeholders will participate meaningfully
   • R: Broad public input may slow development cycles or push toward overly conservative use cases that limit innovation if dominant voices resist new tech.

Score (from 1-3 with, 1 as the best, or n/a) governance actions

Governance Priority

1. Monitoring Governance: for preventing harm in performance data and helping respond to issues. For development of novel biosensor, ongoing surveillance of performance is critical. The action require ongoing performance report (post-market quality data)
2. Targeted Regulatory Standards: to ensure consistent quality before tools reach users. The developmental stage require evidence of safety in realistic biological samples (e.g., urine), and categorize the device based on intended use (environmental screening or clinical decision making)
3. Transparency & Accountability: Publish data of validation results, limitations, and uncertainties.

Resource

• DNAzyme sensors for detection of metal ions in the environment and imaging them in living cells. DOI: 10.1016/j.copbio.2017.03.002
• Heavy Metal Ion Detection Based on Lateral Flow Assay Technology: Principles and Applications. DOI: https://doi.org/10.3390/bios15070438

Assigment (Week 2 Lecture Prep) — DUE BY START OF FEB 10 LECTURE

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?

Answer:

• Error rate of polymerase is 1 : 10^{6 up to 10}8 if there is no correction. It occurs during replocation when DNA Polymerases add wrong nucleotides.
• In compares to the human genome, 1 error per 10^7 nucleotides
• Cell use multiple layers of fidelity mechanisms: Base selection, proofreading (3’–>5’ exonuclease), and mismatch repair.

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?

Answer

• DNA uses four nucleotides (A, T/U, G, C), with each set of three nucleotides (codon). There are 4^3 ways, resulting in around 64 possible codon, but there are only about 24 amino acids because some of the amino acids are coded by more than one codon. Most protein containing about 300 amino acids, which requires 300 codons including start and stop codon.
• Not all sequences code for protein because there are possibilities where cells prefer certain codons, the variety of mRNA structure, and constraints that affect gene expression

Source: https://courses.cs.duke.edu/fall13/compsci101/assign/02_bioinformatics/

Dr. LeProust

1. What’s the most commonly used method for oligo synthesis currently?
2. Why is it difficult to make oligos longer than 200nt via direct synthesis?
3. Why can’t you make a 2000bp gene via direct oligo synthesis?

Answer:

1. Solid-phase Phosphoramidite chemistry method for oligo synthesis
2. Longer oligos increasing number of error rate which reducing the quality and purity
3. Increasing error and low yields, make it unreliable

George Church

1. What are the 10 essential amino acids in all animals and how does this affect your view of the “Lysine Contingency”?
2. (AA:NA and NA:NA codes)] What code would you suggest for AA:AA interactions?
3. [(Advanced students)] Given the one paragraph abstracts for these real 2026 grant programs sketch a response to one of them or devise one of your own:

Answer:

1. Histidine, Isoleucine, Leucine, Lysine, Methionine, Phenylalanine, Threonine, Tryptophan, Valine, Arginine. Lysine COntigency will make animals body depends on lysine being available in the right amount,when lysine is limiting the body can’t build protein effectively, slower growth, and weakening immune function.
2. Lys (K) - Arg (R), Asp (D) - Glu (E), Ser (S) - Thr (T) ???

HTGAA Website DONE ❤️