Life and Environmental Sciences
Reverse transcriptase (RT) PCR
ple: Pale (ple) is a tyrosine hydroxylase that functions in the first and rate-limiting step of dopamine synthesis
Manganese is prevalent in Montana, and Manganese toxicity symptomatically resembles Parkinsons disease. The goal of this project was to determine whether exposure to high levels of Manganese affects the Dopamine synthesis pathway. For our experiment, we chose to measure the expression of the ple gene in Drosophila melanogaster that were exposed to Manganese during development. The ple gene codes for Tyrosine Hydroxylase, an enzyme that functions in the first rate-limiting step of Dopamine synthesis. We hypothesized that an abundance of Manganese would result in decreased expression of ple in larvae and observable motor function deficits among adult flies. To carry out the experiment, flies were randomly assigned either to a control group, cultured under ideal conditions, or a treatment group, cultured in the presence of 0.1mM Manganese Chloride. The experimental group was exposed to Manganese Chloride for 72 hours, after which RNA extraction and Reverse Transcription Polymerase Chain Reaction (RT-PCR) was conducted to compare ple expression. Furthermore, flies were randomly selected to develop into adulthood, after which an assay was performed to compare and quantify motor function in both the control and experimental groups. Because of the oxidative stress Manganese places on the Dopamine synthesis pathway it was predicted that the expression of ple in the experimental groups would be reduced and the developed motor function would be significantly compromised due to the deficiency of Dopamine.
Adair, Brandon; Jones, Tamra E.; and Otto-Hitt, Stefanie, "Manganese Toxicity in the Dopamine Synthesis Pathway in Drosophila melanogaster" (2018). Life and Environmental Sciences Course-based Research Projects. 2.