Week 2 HW: Questions

Homework Questions from Professor Jacobson

Question 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?

DNA polymerase has an error rate on the order of 1 error per 10^{6 bases. Compared to the size of the human genome, about 3 × 10}9 base pairs, this would imply thousands of errors per genome replication. Biology addresses this mismatch by adding layers of correction: polymerases can proofread during synthesis, and additional mismatch repair pathways correct many of the errors that still escape proofreading.

Question 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?

Because the genetic code is degenerate, there are extremely many different DNA sequences that could encode the same average human protein sequence. However, most of these theoretical sequences are not equally functional in real cells. In practice, there are many factors that can strongly affect expression and translation. So although many sequences are valid theorically, only a smaller subset tends to work well in living systems.

Homework Questions from Dr. LeProust

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

The most commonly used method for oligonucleotide synthesis is solid-phase phosphoramidite chemistry. It is a stepwise chemical process where nucleotides are added one at a time on a solid support, which makes it highly automatable and scalable.

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

It is difficult to synthesize oligos longer than roughly 200 nucleotides because errors accumulate at every synthesis cycle. Even if each cycle is “very good,” errors and truncations accumulate with length, so the fraction of full-length, correct oligos drops a lot past ~200 nt.

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

A 2000 bp gene cannot be made reliably by direct chemical synthesis because the cumulative error and truncation rates would make correct full-length molecules extremely unlikely and inefficient. Instead, long genes are typically produced by assembling shorter oligos into larger fragments, followed by cloning and sequence verification to identify correct constructs.

Homework Question from George Church

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

The ten essential amino acids in animals are: histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, valine, arginine