LE3 .A278 2006

2006

Stewart, Don

Acadia University

Bachelor of Science

Honours

Biology

Biology

The objective of this project was to design primers to amplify and obtain DNA sequence data of the Peptidase A (PEP A) gene from shrews. Ultimately these primers could be used to characterize patterns of molecular evolution for this enzyme. Specifically, we are interested in comparing the sequence of a unique allele of PEP A found in the population of S. cinereus on Bon Portage Island compared to alleles of PEP A found in mainland populations of shrews. This is of interest because it has been previously proposed that the unique form of this enzyme may be a molecular adaptation to the unique diet of the shrews on Bon Portage Island. By examining the pattern of synonymous (i.e., neutral or silent) compared to nonsynonymous (i.e., amino acid replacing) substitutions, it would be possible to assess whether positive Darwinian selection is acting on this enzyme. Shrews were trapped on both Bon Portage Island and on the mainland at Sandy Cove, Digby Co., to obtain fresh tissues. These samples were used to extract DNA and RNA. RNA is the starting material used to perform reverse-transcription (RT)-PCR. Successfully amplified bands that were approximately the predicted size for the PEP A product were extracted from agarose gels and purified. These purified PCR products were then inserted into a bacterial plasmid vector and cloned in Escherichia coli cells to create multiple copies of the genetic material so it could be sequenced. A BLAST search of the GenBank database indicated that the amplified PCR product was a gene that codes for an enzyme called carbamoyl phosphate synthetase 1 rather than the desired product, PEP A. Although the sequence of PEP A was not obtained, the sequence of the CPS 1 enzyme might prove useful in the future for phylogenetic studies of the evolutionary relationships of shrews.

The author retains copyright in this thesis. Any substantial copying or any other actions that exceed fair dealing or other exceptions in the Copyright Act require the permission of the author.

https://scholar.acadiau.ca/islandora/object/theses:483