Discipline: Biological Sciences
Subcategory: Cell and Molecular Biology
Kimuel A. Villanova - Baltimore City Community College
High fossil fuel consumption and global warming concerns have encouraged exploration of renewable resources, including non-polar lipids produced by algae that can be converted into biodiesel. A major problem with algae as a source of biodiesel is their slow growth rate under conditions of high lipid production. We are testing the idea that overexpression of genes that increase carbon dioxide (CO2) uptake can increase metabolic function via the Calvin Cycle, leading to improved growth. In this research we are using the model green alga Chlamydomonas reinhardtii to test the effect of overexpressing a Carbon Concentrating Mechanism (CCM) component. The plasma membrane carbonate transporter LCI-1 (Low Carbon Inducible) is responsible for inorganic carbon uptake into the cell. We first generated a nuclear expression vector that contained LCI-1 coding sequence under the control of 5’ and 3’ regulatory sequences (HSP70A-RBSC3 hybrid 5’ UTR + promoter and RBCS2 3’ UTR), but we could not detect LCI-1 protein in any of the transformants. To improve expression, we gene synthesized a bleomycin resistance gene fragment (ble) with a 3’ viral 2A peptide sequence and inserted it directly upstream of the LCI-1 coding region in our first LCI-1 vector. Bleomycin-resistant transformants should produce LCI-1 since ble expression is coupled to the expression of the downstream gene (LCI-1). This construct will be transformed into C. reinhardtii and transformants will be tested via western blot for LCI-1 protein accumulation. Expressing transformants will be analyzed for growth rates in an algal multi-cultivator to compare against the wild type. If this strategy is successful we will apply it to the commonly used industrial alga Chlorella vulgaris for use in biodiesel production.Not Submitted
Funder Acknowledgement(s): Funding for this research is provided by this program is funded by a grant (REM supplement to NSF-EFRI-1332344) from the National Foundation (NSF) Directorate for Engineering (ENG) Office of Emerging Frontiers in Research and Innovation (EFRI). We thank members of the Miller laboratory, including Jose Ortega, Rudolph Park, Erin Pueblo,Jacqueline Rivera, Holly Asbury, Michael Lascola, Jihye Yeon, Alina Buechler, and Sacha Choupa as well as fellow REM interns Sarah Carpe, Charity Ensor, Newton Bosire for their support and technical help.
Faculty Advisor: Stephen Miller, Stmiller@umbc.edu
Role: The Ligation Of LCI-1 and Bleomycin 2a viral peptides. The Prep-work cleaning and mini preps as well as Transformations.