“Using the ABI3 Promoter to Rescue Lethal Editosome Mutations”

Project Summary: 

Plants have a complex RNA-editing mechanism that remedies deleterious T à C mutations by subsequently modifying Cytidine for Uridine (C à U) in organelle transcripts. RNA editing thus acts as a corrective mechanism against harmful mutations and restores the proper encoded protein. Some genes, like OZ2, whose protein productmight be a component of the plant editosome, have not been investigated because they are essential genes. The homozygous mutants do not survive; , and knockdown of these genes by virus gene silencing is sometimes difficult to achieve when the genes are not highly expressed at the RNA level.

To counter this, a conditional promoter, Abscisic Acid-Insensitive3 (ABI3), can be attached in front of the mutant gene, allowing the complementing construct to be expressed during embryogenesis but not to be expressed during seedling development. If the construct carrying the ABI3 promoter and the specific wild-type gene can be properly transformed into the mutant plant via Agrobacterium tumefaciens, then living homozygous mutant progeny can be produced for further study.  During my internship, I have been producing several transformation vectors in which a nuclear-encoded, organelle-targeted protein coding region is placed under the control of the ABI3 promoter.

My Experience:

The most valuable thing I obtained from this internship would be the overall improvement of my ability to create my own experimental design. Although I was operating off a predetermined project, this REU served as an accurate representation for realistic scheduling and setbacks that would be found in my future graduate work. Decisions relating to prioritization of steps during the process fell under my jurisdiction, and this allowed me to manage a more involved role within the project. I was exposed to techniques that were currently evolving and projects that varied across lab specifics and disciplines. Overall, this environment has helped prepare me for my future graduate career in Plant Genetics.