Climate change and global warming threaten the very existence of natural biodiversity by altering ecosystems and distribution of species. This warrants sincere conservation efforts by government and societies to preserve our biodiversity. Conservation and population studies always require precise identification of gender of endangered and elusive species, and this information is very valuable in ecological studies, population genetics, forensic science, ex situ captive breeding programmes, as well as for developing reproductive technologies to boost wild populations. In this chapter, we provide an overview of the importance of gender identification in wild species. We then explain in brief different conventional as well as molecular approaches available for gender identification, with special focus on wild mammals, birds and reptilesReptile. Various methods/markers and procedures used for sex identification are described in detail, along with their advantages and limitations. This will help zoo managers, conservationists, biologists and wildlife researchers who are unaware of all the advanced DNA tools available for sex identification. Overall, this review chapter presents a comprehensive guide to recent advances in molecular sexing, highlighting its strengths and limitations. Research implications and applications in this field are also addressed.