Week 1 HW: Principles and Practices

To make things tidy, I decided to answer most of the questions about the biological engineering application in sections of a separate project idea page.

PLEASE NOTE: on the project page accessible through the link I have been unable to show two images: an image of Van Gogh’s Sunflowers painting and a diagram showing how I’ve scored the actions I’ve suggested. It doesn’t appear to load the image, yet the build log indicates it’s a success. I’m not sure what the problem is but it means you won’t see those two pictures.

Q1. First, describe a biological engineering application or tool you want to develop and why.

See Section 1: Project background

Q2 Next, describe one or more governance/policy goals related to ensuring that this application or tool contributes to an “ethical” future, like ensuring non-malfeasance (preventing harm). Break big goals down into two or more specific sub-goals.

See Section 2: Developing a policy framework to make the project contribute to an ethical future of bioengineering

Q3 Next, describe at least three different potential governance “actions” by considering the four aspects below (Purpose, Design, Assumptions, Risks of “Failure” and “Success”).

See Section 3 Ideation of actions to support policy framework

Q4. Next, score (from 1-3 with, 1 as the best, or n/a) each of your governance actions against your rubric of policy goals.

See 4 Evaluating effectiveness of actions that support policy goals

Q5. Last, drawing upon this scoring, describe which governance option, or combination of options, you would prioritize, and why. Outline any trade-offs you considered as well as assumptions and uncertainties

See 5 Discussions of priorities and assumptions

Lecture Preparation for Week 2

Questions relating to Professor Jacobson’s slides

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?
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 1 The slides and other sources indicate DNA polymerases have a error frequencies of about 10-6 mutations/bp and the human genome has between 3,100,000,000 and 3,200,000,000 base pairs of DNA. ^{1 2} Cells employ various repair mechanisms. Some errors are corrected during replication through a process called proofreading. After replication, mismatch repairs can reduce the rate even further. ³

Answer 2 There are very many ways that proteins could be coded, but most of the combinations do not result in stable three dimensional conformations. The 4th edition of the Molecular Biology of the Cell notes that: “Since each of the 20 amino acids is chemically distinct and each can, in principle, occur at any position in a protein chain, there are 20 × 20 × 20 × 20 = 160,000 different possible polypeptide chains four amino acids long, or 20n different possible polypeptide chains n amino acids long. For a typical protein length of about 300 amino acids, more than 10390 (20300) different polypeptide chains could theoretically be made. This is such an enormous number that to produce just one molecule of each kind would require many more atoms than exist in the universe.

Only a very small fraction of this vast set of conceivable polypeptide chains would adopt a single, stable three-dimensional conformation—by some estimates, less than one in a billion. The vast majority of possible protein molecules could adopt many conformations of roughly equal stability, each conformation having different chemical properties. And yet virtually all proteins present in cells adopt unique and stable conformations.” ⁴

Questions relating to Dr. Le Proust’s slides:

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. According to Twist Bioscience: “Phosphoramidite chemistry is the gold standard method for DNA synthesis that has been used in the industry for almost 35 years. Since its discovery, its simplicity and high efficiency have allowed large volumes of oligonucleotide sequences to be synthesized up to 200 base pairs in length. Currently, it is the only commercially viable chemistry able to provide the volume of DNA required by the synthetic biology market.” ⁵

Answer 2 According to Pichon, it is difficult to synthesise sequences longer than 150 nucleotides because the theoretical yields for larger sequences is low. When solid phase phosphoramidite chemistry is used to make the sequences, the efficiency of coupling between successive oligonucleotides becomes less efficient as the sequence becomes longer. In using this process, the longer the sequence, the more likely that the resulting sequence will contain insertion or deletion errors. ⁶

Answer 3 If two nucleotides are used to make a single base pair, then a 2000 bp gene would require 4000 nucleotides. If synthesising sequences of more than 200 nucleotides is difficult, much more than that would be impractical. Yin describes how “…the state of the art chemical synthesis methods cannot reliably produce oligos longer than 200 nt.” The author explains that for some longer sequences, “…if a sequence contains higher order structures with unusual stability, the PCR assembly method may not function effectively.” ⁷

Question about Prof. Church’s slide #4

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

Answer According to the Cleveland Clinic page on amino acids, the human body needs 20 amino acids to make the all the proteins that make it function properly. However the page and a few other sources only count nine rather than ten essential amino acids. These amino acids are needed by all animals:

• Histidine
• Isoleucine
• Leucine
• Lysine
• Methionine
• Phenylalanine
• Threonine
• Tryptophan
• Valine ⁸

One article names Arginine as a conditionally essential amino acid that is essential in certain life stages or when certain physiological stresses are present. ⁹

If the genetically altered dinosaurs could not produce lysine, they would presumably die before they even reached maturity.