Date of Award

Spring 1992

Project Type

Dissertation

Program or Major

Zoology

Degree Name

Doctor of Philosophy

First Advisor

Directors: Thomas D Kocher

Abstract

This study demonstrates that material preserved in Carnoy's solution (1:3 = glacial acetic acid:alcohol) can be amplified by the polymerase chain reaction for determination of DNA sequence. Because only a single salivary gland from the black fly larva is necessary for DNA extraction, both DNA sequence and chromosome banding data can be obtained from the same individual. This technique reduces potential contamination of the sample by foreign DNA in the gut and/or on the cuticle of the larva and eliminates the possibility of misidentified larvae in the sample. Surveys using the mitochondrial large rRNA gene revealed extensive intraspecific variation in sibling species of the S. venustum and S. verecundum complexes. The occurrence of common mitochondrial DNA (mtDNA) haplotypes in intraspecific samples from different geographic areas suggests the preservation of ancestral polymorphism and/or recent gene flow. Novel mtDNA haplotypes that were unique to individual samples indicate the continuing evolution of the gene or preferential preservation of ancestral polymorphic relicts, or both. The intraspecific variation notwithstanding, mtDNA haplotypes are usually grouped on a conspecific basis. Sibling species of the S. venustum and S. verecundum complexes are well differentiated in the mtDNA genome and can be reliably distinguished by species-specific DNA sequences. For the mitochondrial large rRNA gene of black flies, transition changes are saturated when congeneric or higher hierarchical taxa are compared. The rapid saturation for transition changes might be the result of a bias toward a high proportion of adenine(A) and thymine (T) in the gene. The ability to amplify DNA from single salivary glands of Carnoy's preserved larvae provides the opportunity to conduct DNA, chromosome, and morphological studies on the same individual. The multidisciplinary approach may offer a new perspective on the evolution and ecology of black flies.

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