The spatial distribution of wild germplasm of Podophyllum peltatum L. (American mayapple) has been analyzed using the Geographic Information System (GIS) with the objective to develop a method and a database for evaluation of biotic and abiotic factors influencing drug yield, and to map elite genotypes for propagation and improvement. The field assessment followed a standard procedure including geographical coordinates of each accession, leaf biomass randomly harvested, identification of associate species, collection of herbarium specimen, soil sample and digital pictures of the site. By overlaying morphological and chemical data with geomorphic information, a thematic map was created locating the podophyllotoxin-rich accessions and the uniqueness of each site was recorded for post-collection analysis. This work has enabled the establishment of a database of P. peltatum germplasm in Mississippi with drug yield linked to spatial locations for rational utilization of our natural resources. While this method integrates information of well-characterized diverse in situ P. peltatum germplasm, it might become a strategy for curators to reduce cost for establishing and maintaining ex situ collections since the genetic material is geo-referenced.