Identifying, mapping, and antisensing candidate HP1 sequences in tomato

Abstract

The tomato has been an agriculturally important crop for centuries. For that reason, learning about the fundamental biochemistry and physiology of the plant has been an active research area for decades and has lead to increased crop production. Size, color, nutritional content, herbicide resistance, and insect resistance characteristics have all been successfully manipulated due to the knowledge base established by research and the tools of biotechnology. One main area of research in tomato focuses on fruit ripening. Photo-perception plays a major role in the fruit ripening process and is controlled in part by two high-pigment (HP) genes, HP1 and HP2. When these genes are mutated, the plant appears to have a decrease in hypocotyl length, enlarged cotelydons, and increased anthocyanin levels when germinated in the dark. The fruit appearance of mutants grown in the light includes elevated sucrose, lycopene, chlorophyll and anti-oxidant levels. These characteristics have substantial commercial value. While both mutations are well understood, only the HP2 DNA sequence has been discovered. It was discovered by comparison to a known DNA sequence from Arabidopsis because of the extensive symmetry between Arabidopsis and tomato. I have recently discovered three candidate DNA sequences in tomato that show a high degree of homology to other known negative regulators in the light signal transduction pathway of Arabidopsis. These candidates were discovered by using detailed homology searches based on the Expressed Sequence Tag (EST) database generated by The Institute for Genomics Research (TIGR). After identifying these candidates, I began the basic mapping procedure for identifying general chromosomal location through Restriction Fragment Length Polymorphism (RFLP) mapping. I have also generated antisense mutants of the candidate genes in wild type Lycopersicon esculentum. These mutants are currently in the late stages of tissue culture and may ultimately be used as a tool to clone the HP1 mutant or to identify and clone other genes involved in the light signal transduction pathway of tomato.