The true grasses (Poaceae) comprise one of the largest plant families on earth. The group is peerless in its contribution to global agricultural production and its members dominate many of the world's most important habitats. However, morphological diagnosis of wild grasses is notoriously problematic and it is often impossible in the absence of flowering individuals. The advent of DNA barcoding provided a useful tool to address this problem for larger-scale or longer-term studies but the need for sequencing precludes its use in a field laboratory context or in situations where either funding or time is limited. Here, a chloroplast DNA (cpDNA)-based Cleaved Amplified Polymorphic Sequence (CAPS) system of molecular species diagnosis that has the capacity to address this problem is presented using British grasses as a model. First, PCRs were performed using universal primer pairs targeting 21 regions of the chloroplast genome in authenticated representatives of the 117 grass species from the British Isles, and universal amplification for all loci targeted was demonstrated. Second, 54 restriction enzymes were applied on amplification products generated from all species. There were 10 locus-enzyme combinations (with the highest variation) that had the best diagnostic utility for the 117 grass species.CAPS analysis on 16 representatives of three genera (Calamagrostis, Phleum, and Agrostis) was then used to illustrate the potential utility of the pipeline for establishing a field-laboratory screen of species identity. CAPS DNA barcoding system developed here may have ecological, conservation, and commercial applications. However, it has limited possibilities for intraspecific differentiation due to the highly conserved nature of loci targeted within species.