Grapevine cultivar identification is based mainly on two complementary methodologies: microsatellite (simple sequence repeat (SSR)) DNA analysis and traditional ampelography. Here, we report a direct multiplex PCR approach that allows the simultaneous amplification of 11 SSR loci from crude samples, i.e. bypassing DNA extraction, by using an engineered DNA polymerase improved to tolerate plant PCR inhibitors. Many different plant tissues were successfully amplified: leaf, root, wood, berry flesh and skin, stalk and must, from wine and table grape varieties, and rootstocks. The direct multiplex PCR that we propose is quicker and cheaper than the methodologies used until now, and provides accurate results. Thus, SSR DNA analysis becomes economically more accessible to a larger number of potential users in addition to research institutes.