Submitted by Peggy Lemaux on
New approaches are needed to capture larger atmospheric carbon amounts with the goal of reducing atmospheric CO2, hoping to slow or perhaps even reverse global warming. Plants are a proposed scalable solution, with the possibility to both capture atmospheric carbon through photosynthetic CO2 fixation and store it through carbon sequestration in its roots. Raising the probability of this approach’s success, several studies have shown that photosynthesis is under-achieving and engineered improvements in photosyonthesis demonstrated this. The goal of our project, in collaboration with Professors David Savage in MCB and Krishna Niyogi in PMB, is to accelerate the process of identifying genetic fixes that improve CO2 fixation. Part of achieving this goal will be through the use of a novel screening approach that will lead to identification and prioritization of promising gene overexpression and editing and through searching the literature to discover genes that might have been identified in other crops to have a positive effects on photosynthesis or root biomass..
The ultimate target of our carbon sequestration efforts is Sorghum bicolor, which is the fifth most widely grown cereal crop worldwide. In the U.S. sorghum is primarily used in animal feed and biofuels, but it is an important food crop in other areas. Advantages of sorghum for biofuels is that it uses the more efficient C4 photosynthetic pathway and it is drought- and flood-tolerant, traits important to addressing climate change. A disadvantage of sorghum is that the seed to seed lifecycle is ~4.5 months, making it difficult to make rapid progress in determining the function of genes involved in photosynthetic efficiency and biomass production. Because of this, for initial gene function screens we might use Setaria viridis, a C4 grass with a shorter 7-8 week lifecycle.
Several approaches will be used to identify genes of interest. One approach involves use of sorghum protoplasts to screen genome edits in a high-throughput assay. From those candidate edits, the most promising ones will be further evaluated by creating editing or overexpression constructs to stably transform into sorghum or Setaria. Evaluation of engineered plants will include various physiological and biochemical assays to determine effects of the modifications on photosynthesis and root biomass. Another approach to identifying candidate genes for improvement was to screen the literature to identify traits that appear to have conferred improved photosynthesis or increased biomass on plants, sorghum or otherwise. Through this process, to date we have focused on two genes, Raf1 (rubisco accumulation factor) and Zmm28 (MADS-box transcription factor). A construct was created that will lead to overexpression of Raf1, introduced into the RTx430 sorghum genotype and T0 transgenic plants generated. Another construct is being created to overexpress Zmm28.
Essential to these efforts are effective methods to transform sorghum. From a 1-3% transformation efficiency using a classical immature embryo transformation method, we moved to a nearly 50% efficiency utilizing the developmental genes, Bbm and Wus (Aregawi, Shen et al. 2021. Plant Biotech J doi: 10.1111/pbi.13754). Even though transformation efficiency is high, editing and knockout efficiency in the T0 generation are 22 and 17 % respectively. We plan to improve this by modifying various components of the previously used construct. Different target genes to improve C4 photosynthesis and root system will be edited using this approach. Improving knockout and editing efficiencies will be vital to easily generating sufficient numbers of genome-edited plants.
The duties and goals of the student will depend on their skill level. The student might care for plants, plant seeds, harvest seeds, perform in vitro tissue culture and transformation for overexpression/editing and conduct biochemical and molecular studies on the transformed plants. The student may also be involved in preparing materials for the various analyses that will be performed. The student may also care for plants in growth chambers and the greenhouse, collecting experimental materials and data when necessary. The student will work directly with the Staff Research Associate, postdoctoral fellows, other undergraduates and the Principal Investigator. Students will participate in lab meetings and be expected to contribute to presentations and publications, as appropriate. The time commitment required will be negotiated with the student.
Enthusiasm for research is necessary; previous laboratory experience outside the classroom is desirable. Care-to-detail and commitment to scheduled work times are critical.