These data represent nuclear microsatellite data collected from four riparian plant species that occur in and around Grand Canyon National Park: Populus fremontii (POFR), Salix gooddingii (SAGO), Salix exigua (SAEX), and Prosopis glandulosa (PRGL). These data were collected for population genetic analysis to help inform native plant material development in Grand Canyon National Park. Leaf samples were collected at sites spanning 470 km of the Colorado River between Glen Canyon Dam and Lake Mead and in tributaries. Known revegetation areas were not sampled. We aimed to collect leaf tissue from at least 15 individuals at each sample site. If there were fewer than 15 individuals per species at a site, leaf tissue was collected from every individual. Leaves were immediately dried and stored in silica gel. For P. fremontii, S. gooddingii, and P. glandulosa, total genomic DNA was extracted from dried leaf tissue using a high-molecular weight protocol with modifications. For S. exigua, DNeasy Plant Minikits were used. We amplified 9, 9, 11, and 13 loci for P. fremontii, S. gooddingii, S. exigua, and P. glandulosa, respectively. All loci were amplified using polymerase chain reaction (PCR) and fragment analysis processed on an ABI 3730xl Genetic Analyzer with GeneScan LIZ500 internal size standard. Allele fragment sizes were scored using GeneMarker v2.2.0. Loci that were missing more than 5% of values, were not polymorphic, or could not be reliably scored were omitted. Nine loci were included for P. fremontii, six for S. gooddingii, eight for P. glandulosa, and nine for S. exigua. All species are diploid, so each microsatellite locus has two values. Associated with the microsatellite data are labels delineating what collection site each individual came from, which genetic group each individual was assigned to during statistical analyses, and information about those sites. Specifically, sites are noted as being on the Colorado River or on a tributary to it, and inside or outside of the canyon rims.