Joshua Bradshaw
Joshua Bradshaw headshot
Year: 2023
Faculty Advisor: James Giovannoni

Carotenoid analysis of gene-edited rin mutant tomatoes

Tomato (Solanum lycopersicum) serves as a model system to study ripening in fleshy fruit due to its fast rate of growth (3-4 months) and ease of genetic transformation. The rin (ripening inhibitor) mutation is naturally occurring in tomato, and results in a fused chimeric message between the RIN gene (transcription factor known to regulate ripening processes) and the adjacent MC gene (transcription factor involved in the development of plant inflorescence) on the chromosome. In this fused message, the 8th exon in RIN is deleted and part of the promoter in MC is deleted. The result of this mutation is fruit that fails to produce ethylene (a ripening hormone), never synthesizes carotenoids (responsible for red/orange/yellow color), doesn’t soften, and doesn’t develop a ripe aroma. To determine why the rin mutant tomatoes never ripen, CRISPR Cas-9 was used to effectively knock different parts of the RIN/MC message, and observe the phenotypes of the CRISPR lines. Carotenoid levels were measured in each CRISPR line to find the difference between carotenoid content in CRISPR lines compared to wild type tomatoes as well as the rin mutant, with the goal of discovering why the mutant tomato doesn’t ripen. With this knowledge, tomatoes that ripen slower, ship easier, and have a longer shelf life, can be used to feed the world.

I have had a great experience in the Giovannoni lab this summer. I have improved my ability to run PCR and gels, and learned new skills like gel purification, DNA extraction, and carotenoid extraction. I really enjoyed attending the weekly presentations and symposiums, and working in a hands-on lab environment. I also enjoyed getting to know my fellow high school interns, as well as the undergraduate interns. I would like to thank Jim Giovannoni for giving me the opportunity to work in his lab this summer. I also especially want to thank my mentor, Julia Vrebalov, for answering endless questions, guiding me through my research, and being patient with me and my fellow interns throughout the learning experience.