Changes in the structure, content and morphology of chromosomes accumulate over evolutionary time and contribute to cell, developmental and organismal biology. The axolotl (Ambystoma mexicanum) is an important model for studying these changes because: 1) it provides important phylogenetic perspective for reconstructing the evolution of vertebrate genomes and amphibian karyotypes, 2) its genome has evolved to a large size (~10X larger than human) but has maintained gene orders, and 3) it possesses potentially young sex chromosomes that have not undergone extensive differentiation in the structure that is typical of many other vertebrate sex chromosomes (e.g. mammalian XY chromosomes and avian ZW chromosomes). Early chromosomal studies were performed through cytogenetics, but more recent methods involving next generation sequencing and comparative genomics can reveal new information. Due to the large size and inherent complexity of the axolotl genome, multiple approaches are needed to cultivate the genomic and molecular resources essential for expanding its utility in modern scientific inquiries. This dissertation describes our efforts to improve the genomic and molecular resources for the axolotl and other salamanders, with the aim of better understanding the events that have driven the evolution of vertebrate (and amphibian) chromosomes. First, I review our current state of knowledge with respect to genome and karyotype evolution in the amphibians, present a case for studying sex chromosome evolution in the axolotl, and discuss solutions for performing analyses of large vertebrate genomes. In the second chapter, I present a study that resulted in the optimization of methods for the capture and sequencing of individual chromosomes and demonstrate the utility of the approach in improving the existing Ambystoma linkage map and generating targeted assemblies of individual chromosomes. In the third chapter, I present a published work that focuses on using this approach to characterize the two smallest chromosomes and provides an initial characterization of the huge axolotl genome. In the fourth chapter, I present another study that details the development of a dense linkage map for a newt, Notophthalmus viridescens, and its use in comparative analyses, including the discovery of a specific chromosomal fusion event in Ambystoma at the site of a major effect quantitative trait locus for metamorphic timing. I then describe the characterization of the relatively undifferentiated axolotl sex chromosomes, identification of a tiny sex-specific (W-linked) region, and a strong candidate for the axolotl sex-determining gene. Finally, I provide a brief discussion that recapitulates the main findings of each study, their utility in current studies, and future research directions. The research in this dissertation has enriched this important model with genomic and molecular resources that enhance its use in modern scientific research. The information provided from evolutionary studies in axolotl chromosomes shed critical light on vertebrate genome and chromosome evolution, specifically among amphibians, an underrepresented vertebrate clade in genomics, and in homomorphic sex chromosomes, which have been largely unstudied in amphibians.