Abstract: Biodiversity provides the basic environmental and material conditions for the survival and development of human society. With the increasingly prominent problems of biodiversity, there are higher requirements for the research and protection of biodiversity. As the basic work of biodiversity research and conservation, it is particularly more important to improve the accuracy and convenience of field investigation. Two-step outdoor assistant App is a professional outdoor software that can be used on mobile phones and tablets, with offline map downloading and navigation, route design, track recording, interest points marking, data sharing and other functions, which can provide great convenience and reliability for biodiversity investigation. The paper introduced the realization of the related functions of the two-step outdoor assistant App software in biodiversity investigation, and compared it with three outdoor softwares of the same type, such as Lotoo, Ovi map, Foooooot map, showing the accuracy and convenience of the two-step outdoor assistant App in biodiversity survey, so as to provide technical and methodological references for biodiversity survey and research.