For the 2018 iGEM competition, we plan to create a plasmid vector with another strain of yeast, Pichia pastoris. Plasmid vectors are invaluable tools for synthetic biologists and the broader scientific community. For those who work with yeast, one particularly useful vector is the centromeric plasmid: a vector that, by essentially mimicking a yeast chromosome, is able to provide both the flexibility of replicating plasmids and the stability of integrating plasmids.

Organisms often do not retain plasmids once they have been transformed. Plasmid loss occurs when cells undergo replication. During mitosis, microtubules attach to the centromere of DNA and distribute it evenly among the daughter cells. If we were to insert a centromeric sequence into a plasmid, the microtubules would attach to this region of DNA. As a result, the plasmid would be distributed much more equally (and without loss) among the daughter cells in mitosis.

While there are multiple centromere plasmid options for Saccharomyces cerevisiae, the scientific community has yet to design a yeast centromere series for the other widely-used yeast strain, Pichia pastoris. In response, UChicago Genehackers has set out to design the first yeast centromeric plasmid for Pichia pastoris.

We will identify the minimal, essential region of the Pichia centromere that, when incorporated into a plasmid backbone, will allow the plasmid to mimic a Pichia chromosome during replication. This region can then be incorporated into Pichia backbones to convert them into centromere plasmids, which transform with high efficiency, have low and constant copy numbers, and are stably maintained without integration. Ultimately, we will contribute an important foundational tool for flexible, creative, and impactful cloning in yeast. If successful, the centromeric plasmids will serve a multitude of purposes in the bioengineering and industrial fields.