The size of a natural population may be referred to the number of accessions or the areas where the individuals are collected. However, this concept has never been well defined in theory or practice. The genetic diversity between one natural population of wild barley and barley varieties from different countries were investigated using two sets of independent molecular markers: SSR and intron-splice junction primer-based PCR. The result suggests that the level of genetic diversity of one natural population of wild barley is much higher than that of a collection of barley varieties sampled from various locations around the world. Comparable data were found in an extension of this project with a d another wild barley population and more sampling of barley varieties from various parts of the world, which support this conclusion. It is well known that domestication and selection have resulted in drastic impoverishment of the gene pools of cultivars. Still, it is important to note that the genetic diversity among barley varieties in the entire world, as represented by this sampling, was found to be much lower than that of a wild barley population from a site in the Jordan Valley. Our results put forward a range of implications and ramifications to the theory and practice for plant germplasm collecting, conservation, and storage.