Obtaining reliable methods for estimating genetic diversity of large populations, especially those of non-model organisms, can provide versatile tools in various research fields including genetic/evolutionary rescue and biodiversity effects on ecosystem functioning. Recent technological advancements in next-generation sequencing have significantly improved the quality of genomic analyses while making high-throughput genetic sequencing more time- and cost-effective. This study assesses the viability and extent of the pool-GBS method in estimating population genetic diversity of the Japanese mountain birch (Betula ermanii). This fast-growing pioneer tree species is dominant across subalpine/alpine forest ecosystems in Hokkaido, Japan. Young leaf tissues of two separate sapling populations were collected from different locations within the Uryu Experimental Forest in Hokkaido. Genomic DNA was digested using PstI and MspI restriction enzymes, and adapters were ligated with T4 ligase. Individual and pooled (i.e. 12, 18, 24, 30, and 36 individuals) DNA samples were barcoded with indexed primers. For library assembly, DNA fragments of 350-800bp were selected. The samples were then sequenced with MiSeq using a PhiX control. The results of genetic diversity analyses assessing population allele frequency, heterozygosity, and genetic differentiation were compared between pooled and non-pooled populations to determine reproducibility.