Honors Capstone Project
Date of Submission
John Belote, Professor
Melissa Pepling, Associate Professor
Arts and Science
Capstone Prize Winner
Won Capstone Funding
Sciences and Engineering
Speciation occurs when two populations of a species can no longer reproduce, either because of (1) pre-mating reproductive isolation, for example, populations having non-overlapping habitats, or evolving different courtship behaviors, or (2) post-mating pre-zygotic (PMPZ) isolation, where mechanical (e.g., incompatible differences in genitalia), or gametic (e.g., sperm and egg become incompatible) differences renders the strains unable to produce offspring. Previous work has suggested that when incipient species begin to experience reproductive isolation, a phenomenon called “reinforcement” can accelerate the process, by the rapid evolution of new pre- mating or PMPZ barriers. While the occurrence of reinforcement has been studied for many years, not much is known about how rapidly natural selection can create new reproductive barriers, or what the actual mechanisms are that are likely to arise [e.g., pre-mating mechanisms like changes in courtship behavior, or PMPZ mechanisms like differential sperm storage or use by the female (cryptic female choice)]. My project focuses on using genetic engineering to create populations of genetically incompatible strains of Drosophila melanogaster, which then will be studied to understand the exact mechanisms whereby such strains might rapidly diverge from each other due to reinforcement. My research has used recombinant DNA methods to create two complex synthetic alleles (A and B), each consisting of four components. (1) The Prot-B RFP or Prot-B GFP sequences that encode sperm specific proteins tagged with red fluorescent proteins (RFP) or green fluorescent protein (GFP) for the clear distinction of A and B sperm within a female’s seminal receptacle; (2) the 3xP3 RFP and 3xP3 GFP cassettes that result in eye-specific expression of RFP and GFP to allow easy identification of which allele each fly carries; (3) the attB sequence that allows these constructs to be inserted into a specific chromosomal site, using the phiC31 integrase system for site-specific transformation; and (4) one of the two components of the GAL4/UAS system for targeted gene expression. The A allele carries the yeast transcriptional activator protein gene, GAL4, under the control of a constitutive promoter (i.e., the Pros25 proteasome gene promoter). The B allele carries the dominant lethal gene UAS-Poly-Q108 that is activated by GAL4. Thus, while either allele by itself is harmless, when both are present, such as in AB hybrids, the activation of UAS-PolyQ108 leads to 100% pupal lethality. These newly created incompatible A and B populations will be used for long-term experimental evolution studies. In these experiments, flies from both strains will be maintained together in population cages where they can freely mate and reproduce for many generations. Because the only productive matings will be between A x A and B x B, due to the hybrid incompatibility, there will be selective pressure to quickly evolve additional reproductive isolation mechanisms, which will then be identified and analysed in detail. If successful, this will confirm the concept of reinforcement, and furthermore give us clues about specific reproductive isolation mechanisms that are likely to quickly evolve.
Poplawska, Maria, "Developing an Experimental System for Studying Early Events in Speciation in Drosophila" (2013). Syracuse University Honors Program Capstone Projects. 68.
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