Event Title

: Investigang plastome expression of Rubisco Activase in Chlamydomonas reinhardtii as a platform for directed evolution

Advisor(s)

Dr. Sarah Stainbrook; Washington University of St. Louis

Discipline

Biology

Start Date

21-4-2021 10:05 AM

End Date

21-4-2021 10:20 AM

Abstract

Rising temperatures due to global warming spur concerns over whether plant life will be able to feed the rapidly growing global population. One solution is to improve the resilience of plants to climate change through genetic modification. Rubisco activase (RCA) is a particularly important target because it loses efficiency rapidly at high temperatures, impairing photosynthesis. Our goal is to engineer a system for expressing Rubisco Activase variants from the chloroplast genome in Chlamydomonas reinhardtii. We can characterize the effects of these variants on assimilation and cell growth at various temperatures. Ultimately, this system will enable directed evolution of RCA isoforms for improved thermotolerance. To do this, we have built plasmids with several different promoters using Golden Gate cloning, integrated them into the Chlamydomonas plastome, and characterized gene expression from each promoter by measuring the expression of the mVenus fluorescent protein. In this work we compare the growth of RCA-knockout strains vs. the wild-type in multiple media types. We plan to use the data of this experiment in next year’s research, where we will investigate whether different RCA types can be used to characterize heat tolerance of algal photosynthesis in order to improve the heat tolerance of RCA via directed evolution.

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Apr 21st, 10:05 AM Apr 21st, 10:20 AM

: Investigang plastome expression of Rubisco Activase in Chlamydomonas reinhardtii as a platform for directed evolution

Rising temperatures due to global warming spur concerns over whether plant life will be able to feed the rapidly growing global population. One solution is to improve the resilience of plants to climate change through genetic modification. Rubisco activase (RCA) is a particularly important target because it loses efficiency rapidly at high temperatures, impairing photosynthesis. Our goal is to engineer a system for expressing Rubisco Activase variants from the chloroplast genome in Chlamydomonas reinhardtii. We can characterize the effects of these variants on assimilation and cell growth at various temperatures. Ultimately, this system will enable directed evolution of RCA isoforms for improved thermotolerance. To do this, we have built plasmids with several different promoters using Golden Gate cloning, integrated them into the Chlamydomonas plastome, and characterized gene expression from each promoter by measuring the expression of the mVenus fluorescent protein. In this work we compare the growth of RCA-knockout strains vs. the wild-type in multiple media types. We plan to use the data of this experiment in next year’s research, where we will investigate whether different RCA types can be used to characterize heat tolerance of algal photosynthesis in order to improve the heat tolerance of RCA via directed evolution.