FRIDAY PAPER PRESENTATIONS

Georgia Journal of Science, 2006

*Denotes student presentation

**Denotes student in-progress research

Section I: Biological Sciences

B-1500

Mark S. Davis and Terry D. Schwaner, presiding

1:30 MUTAGENESIS OF THE SEC15 SUBUNIT OF THE EXOCYST COMPLEX IN YEAST**, Daniel B. Price*, Chevenne Simmons*, Michael Gleason and Y. Ellen France, Georgia College & State University, Milledgeuille, GA 31061. The exocyst complex is required to properly target secretory vesicles to the daughter cell of the budding yeast in Saccharomyces cerevisiae. This project is designed to create functionally significant mutations in the SEC15 subunit of the exocyst complex. A PCR product amplified from the SEC15 gene will be restricted and the C-terminal encoding region (SEC15C) will be spliced into a pPG5 vector. This plasmid (SEC?5C-pPG) will after its transformation into XL1-Red Escherichia coli cells (Stratagene) undergo random base-substitution mutagenesis. A heterogeneous mix of mutagenized plasmids (SEC15^sup m^-pPG) will be isolated and restricted. This mix of mutagenized restriction fragments (SEC15^sup m^-pPG) will then be spliced into a similarly digested SEC15-pPG5 construct to replace its wild-type C-terminal encoding regions. The resulting pool of mutant constructs (SEC15N-sec15C^sup m^-pPG5) will then be restricted and spliced into a pool of similarly restricted LEU2-based yeast integration plasmids. After homologous recombination of the SEC15N-sec15C^sup m^-LEU2 plasmid into yeast, bio-assay will be used to identify useful mutants. This research is supported in part by a GC&SU Faculty Research Grant and the GC&SU Foundation.

1:45 PHENOTYPIC SCREENING OF MUTAGENIZED SEC15 PROTEIN, AN ESSENTIAL SUBUNIT OF YEAST SECRETORY EXOCYST COMPLEX", David Nix*, Munis Lukman, Michael Gleason and Y. Ellen France, Georgia College & State University, Milledgeville, GA 31061. Sec15 is an essential protein subunit of the exocyst complex required for proper targeting of secretory vesicles from the parent cell to the budding daughter cell membrane in the yeast, Saccharomyces cerevisiae. The C-terminal region of secl5 protein will be randomly mutagenized (sec15Cm) and then screened for characteristic phenotypes indicative of secretory defect in yeast. A haploid sec 15Δ Ieu2 ura3 yeast strain carrying a 2µ sec15-URA3 plasmid will be transformed with a SEC15N-sec15Cm-LEU2 integration plasmid that will recombine with the defective leu2 chromosomal locus. The wild-type 2µ SEC15-URA3 plasmid will be shuffled out of the yeast by counter selection with 5'-FOA. Individual clones of leu^sup ^ ura^sup -^ cells each containing one of several possible random secl5-Cm mutations will then be bio-assayed for growth at 25, 30, 34 and 37°C. Yeast clones with temperature- sensitive growth will be further screened for altered morphological, localization, solubility and biochemical phenotypes. Clones exhibiting characteristic phenotypic changes will undergo PCR-based sequencing to discover the physical nature of each Sec15 mutation. The development of new Sec15 mutants should further studies intended to elucidate its role in exocyst function. This research is supported in part by a GC&SU Faculty Research Grant and the GC&SU Foundation.

2:00 THE EFFECT OF ALL-TRANS RETINOIC ACID ON CARDIAC MORPHOGENESIS IN ORYZIAS LATIPES: A LOOK INTO GENE EXPRESSION PATTERNS**, Uschi Auguste* and Holly Boettger-Tong, Wesleyan College, Macon GA 31210. Retinoic acid is a potent derivative of Vitamin A that affects vertebrate physiological processes such as cell growth and differentiation, and morphogenesis. Retinoic-acid mediated gene activation is important for normal vertebrate development; however, when embryonic exposure to retinoids is higher than normal, developmental anomalies occur. This investigation focuses on the effect of supplying all-trans retinoic acid (ATRA) to neurula Oryzias latipes (Japanese Medaka) embryos, with specific interest in changes in cardiac morphology and changes in gene expression patterns, specifically in genes pertinent to cardiogenesis, vasculogenesis, and apoptosis. Extracted messenger RNA from control and ATRA treated animals will be compared using a 4 x 2K custom DNA microarray cassette containing approximately 200 genes. Genes represented on this microarray have been implicated in the control of cardiogenesis, vasculogenesis, and cell death pathways in Medaka and other organisms. We have also added zebrafish and mouse sequences to this array to test the hypothesis that cross-species hybridization between Medaka mRNA and orthologous genes is possible. Using this microarray, we hope to identify patterns of gene expression that have been altered by all-trans retinoic acid treatment in developing Medaka embryos.

2:15 CLASSICAL AND MOLECULAR BIOLOGY APPROACHES FOR CHARACTERIZATION OF MICROBIAL COMMUNITIES INDIGENOUS TO CLAYS**, Hannah J. Hoehn*, Glenda Kohlhagen, and Andrei L. Barkovskii, Georgia College & State University, Milledgeville, GA 31061. clays comprise a substantial portion of terrestrial and aquatic solids including soils and sediments. Currently, microbial communities of clays are virtually unknown. Given that microbial community characterization is heavily based on DNA-dependant methods, this lack of knowledge is mainly due to difficulty with DNA isolation from clays. We attempted to characterize the microbial community composition of typical Georgia clays via a customized isolation protocol allowing for the isolation of freely-dispersed and particle-associated clay microorganisms. The completeness of isolation was tested through direct DNA extraction from clays. No DNA was obtained from clays or from previously isolated microbial fractions, although direct plate counts demonstrated a high abundance of microorganisms. We also failed to identify a DNA isolation method appropriate for clays or clay-contaminated microbial communities in the literature. The current presentation provides an analysis of various custom-designed methods and their effectiveness towards DNA isolation from clay-containing media. These studies are supported by two grants from IMERYS located in Sandersville, GA.