Reverse Transcription-Polymerase Chain Reaction (RT-PCR) analysis of Histidine Decarboxylase Transcription in Drosophila melanogaster
Histamine is a biogenic amine that is used as a neurotransmitter in both vertebrates and invertebrates. Histidine decarboxylase (HDC) is the enzyme that synthesizes histamine using histidine as a substrate. Mutations in the HDC gene of Drosophila melanogaster cause blindness and other functional defects in the fly. Sequence analysis of 5’ and 3’ cDNA ends identified through 5' and 3' RACE (Rapid Amplification of cDNA Ends) has suggested that there are alternative transcript initiation and polyadenylation sites for the histidine decarboxylase transcription unit. Using these findings, as well as mRNA sequence information previously obtained, primers were developed to analyze the alternative HDC transcripts. Messenger RNA was isolated from fly heads and used as a template for RT-PCR analysis. The results confirm the existence of multiple transcripts with initiation and splicing sites that were previously not identified. RT-PCR results also provide evidence to support the presence of a second polyadenylation site at the end of the HDC mRNA. Analysis of these alternative transcripts, using RT-PCR and Q-PCR approaches, will provide further insight into transcriptional regulation of HDC by examining the levels of the transcript and tissue specificity.
Faculty Mentor: Martin Burg
Dan presented at the National Drosophila Research Conference April 2-6, 2008 in San Diego, CA.
Page last modified July 14, 2009