Rapid advances in DNA sequencing technologies made it cost-effective to get high-throughput genomic sequence data. Applications in whole genome resequencing presented new approaches to investigate natural variation and adaptation of species. The genus Populus in the Salicaceae family includes economically and ecologically critical species that are dioecious, wind-pollinated, and predominantly distributed in river ecosystems. Poplar serves as a model for the research on the evolutionary genomics of forest tree species. In this study, we aimed to present an example of a comprehensive SNP calling pipeline, demonstrating its application in elucidating genetic diversity and population structure from whole genome resequencing data. The results discovered genetic variants from whole genome resequencing data of poplar species. The pipeline involved sequence preprocessing, alignment to the reference genome, and variant calling.