Optimization of PCR conditions to amplify microsatellite loci in the bunchgrass lizard (Sceloporus slevini) genomic DNA

<p>Abstract</p> <p>Background</p> <p>Microsatellites, also called Simple Sequence Repeats (SSRs), repetitions of nucleotide motifs of 1-5 bases, are currently the markers of choice due to their abundant distribution in the genomes, and suitability for high-throughput analysis. A total of five different primer pairs were optimized for polymerase chain reaction (PCR) to amplify microsatellite loci in total genomic DNA of bunchgrass lizards (<it>Sceloporus slevini</it>) collected from three sites in southeastern Arizona; the Sonoita Plain, Chiricahua Mountains and Huachuca Mountains.</p> <p>Findings</p> <p>The primers used for current investigation were originally designed for the Eastern Fence Lizard (<it>Sceloporus undulatus</it>). Five primer pairs were selected based on annealing temperatures for optimizing the PCR conditions to amplify with bunchgrass lizards. Different concentrations of DNA and annealing temperature were optimized. While keeping other reagents constant, a DNA concentration, 37.5 ng in the final reaction volume and PCR conditions of an initial denaturation of 94°C for five minutes, an annealing temperature of 55°C and final extension of 72°C for four minutes gave the best amplification for all the primer pairs.</p> <p>Conclusions</p> <p>Modifying the standard protocol for annealing temperatures and final extension time increases the success of cross amplification of specific microsatellite loci in the bunchgrass lizard. A loading volume of 5 ul DNA at a concentration of 10 ng/ul and a 2% agarose for gel electrophoresis were observed the best for cross amplification of selected five primer pairs on bunch grass lizard.</p> <p>Trial Registration</p> <p>The research was conducted with Arizona Game and Fish Department scientific collecting permits SP565256, SP657407 & SP749119 to Dr. Christian A d'Orgeix.</p

Dietary niche partitioning of three Sky Island Sceloporus lizards as revealed through DNA metabarcoding

Abstract Lizard diets are highly diverse and have contributed to the diversification, biogeographical distributions, and evolution of novel traits across this global radiation. Many parts of a lizard's ecology—including habitat preferences, foraging modes, predation risks, interspecific competition, and thermal constraints, among others—interact to shape diets, and dietary niche partitioning simultaneously contributes to co‐occurrence within communities. We used DNA metabarcoding of fecal samples to identify prey items in the diets of three sympatric Sceloporus lizards in the Madrean Sky Islands of Arizona, USA. We found evidence for dietary niche partitioning between interacting species concomitant with their respective ecologies. We also compared diet composition between populations to understand how conserved or plastic species' diets are between different environments. Our findings suggest that habitat generalists are also diet generalists in this system, while the same may be true for specialists. The identification of prey items to much lower taxonomic levels than previously documented further reveals hidden diversity in the diets of these species and underscores the utility of metabarcoding for understanding the full complexity of lizard diets