Xanthocyparis vietnamensis is an endangered species that is currently restricted to karst montane areas in southwestern China and Vietnam. This rare conifer was first recorded in 2002 from northern Vietnam and then in 2013 from Guangxi, China, yet little is known about its genetic diversity and population structure. We developed expressed sequence tag microsatellite markers (EST-SSR) for X. vietnamensis. Illumina HiSeq data were used to reconstruct the transcriptome of this species by de novo assembly. Based on this transcriptome, 20 SSR markers were designed and successfully amplified, and no loci with excessive null allele was detected. These polymorphic loci were then employed to access the genetic diversity and population structure of X. vietnamensis. As well, they could be useful in promoting sustainable forest management strategies for this species in the vulnerable karst ecosystem.