AbstractOmphalogramma soulieiFranch. is an endangered perennial herb only distributed in alpine areas of SW China. ISSR markers were applied to determine the genetic variation and genetic structure of 60 individuals of three populations ofO. soulieiin NW Yunnan, China. The genetic diversity at the species level is low withP=42.5% (percentage of polymorphic bands) andHsp=0.1762 (total genetic diversity). However, a high level of genetic differentiation among populations was detected based on different measures (Nei's genetic diversity analysis:Gst=0.6038; AMOVA analysis:Fst=0.6797). Low level of genetic diversity within populations and significant genetic differentiation among populations might be due to the mixed mating system in which xenogamy predominated and autogamy played an assistant role inO. souliei.The genetic drift due to small population size and limited current gene flow also resulted in significant genetic differentiation. The assessment of genetic variation and differentiation of the endangered species provides important information for conservation on a genetic basis. Conservation strategies for this rare endemic species are proposed.