Genome Engineering

19th of October - George Church (Harvard) and John Glass (JCVI)

Class slides Recordings

Please answer these questions

1. Craig Venter has stated he will establish an IGEM style competition for the most innovative use of the JCVI minimal bacterial cell, JCVI-syn3.0. The JCVI built the organism to be used as a platform to investigate the first principles of cellular life. Please describe how you would use JCVI-syn3.0 to investigate basic cellular biology or to use the organism to learn how to make a better cell for some research, medical, or industrial purpose. Do not get stymied by technical details involving how you would genome engineer JCVI-syn3.0. Explain the concept and experimental rationale.

2. In previous classes, George Church has talked about work done in his lab to do grand scale genome engineering of E. coli to alter its genetic code. While there are similarities with how his lab engineers E. coli with how the JCVI engineers mycoplasmas, what is the fundamental difference or differences?

3. In the Science paper “Design and synthesis of a minimal bacterial genome,” the JCVI showed the figure below, which is the first step in an effort to rationally reorganize the minimal cell genome. As noted in the paper, when we reorganized the genome, which included separating genes in operons, as needed we placed genes behind transcriptional promoters that controlled expression of genes deleted to build minimized segment 2. We built a genome that had a reorganized segment 2, but with the other 7 segments in with their original gene order. As noted in the paper, that cell grew normally.

In later experiments, the JCVI used the same strategy to design and build reorganized versions of the other 7 minimized segments and tested them as genomes that were 1/8th reorganized and 7/8ths not reorganized. None of those 7 genomes resulted in viable cells. What is your hypothesis as to why this happened?

4. The JCVI recently announced that it was now minimizing and reorganizing the genome of the fastest growing eukaryote, a yeast called Kluyveromyces marxianus. The goal will be to make an alternative to Saccharomyces cerevisiae that grows faster, and can be grown at higher temperature, and that would be a better platform for the kinds of biotechnology people currently use yeast for. Based on the “genome engineering lecture” in HTGAA and the Science paper “Design and synthesis of a minimal bacterial genome” (assigned as class reading), how would you go about minimizing and rationally reorganizing the K. marxianus? What would be different about your approach relative to how the JCVI minimized Mycoplasma mycoides to produce JCVI-syn3.0?