This project will attempt to identify, describe, compare, and contrast the sex determination systems of two different groups of reptiles, pit vipers (subfamily Crotalinae) and tortoises (family Testudinidae). Pit vipers have heterogametic females (ZW, males being ZZ) which determine the sex of their offspring by classical sex chromosome segregation (GSD = Genetic Sex Determination). Tortoises lack sex chromosomes. They are believed to resolve sex through Environmental Sex Determination (ESD), though this has only been demonstrated in one of nearly fifty species (Testudo graeca, Pieau, 1975). NORs and C and G-banded karyotypes will be obtained for all genera in both families (of subfamilies). A new pit viper phylogeny, based upon separate sets of karyotypic, allozyme, DNA-DNA hybridization, visceral topography and karyotypes will be constructed for this widespread and extremely dangerous group of poisonous snakes. An accurate phylogeny will guide toxicologists and immunologists in their search for antibodies effective against the venom components of close relatives as well as pharmacologists in their search for related sources of venom fractions valuable to medicine. Continuous monitoring of natural nests of desert tortoise (Gopherus Agassizii) eggs at Ft. Irwin, California, will provide new information about ambient temperatures fluctuation, relative humidities and gas partial pressures. This information will make possible novel experiments subjecting captive tortoise eggs to variations (both degree and duration) in these same parameters which may produce skewed sex ratios in hatchlings. Morphometric techniques for non-invasive sexing of neonates are proposed using three dimensional fractal geometry. Collaborative studies with MBRS PI John Roberts will include Souther blot hybridization and subsequent in situ hybridization of the mammalian male Sry gene on snake and tortoise genomic DNA, and periodic monitoring of RNA and protein synthesis during tortoise embryogenesis. These changes in gross chromosome morphology, and help explain the selective pressures producing the phylogenetic differences within both groups. In this way, both universal and taxon specific elements in the genetics of sex determination might be resolved.
