The cytochrome c oxidase subunit I (COI) gene of the mitochondrial DNA was sequenced in eight holocyclic monoecious aphids (Hemiptera: Aphididae) that occur on wheat, Triticum aestivum L.; barley, Hordeum vulgare L.; oat, Avena sativa L.; and sorghum, Sorghum bicolor (L.) Moench in the United States. The first 640 bp of the 5′ end were considered as a DNA barcoding technique for species identification. DNA barcoding successfully differentiated Schizaphis graminum (Rondani), Diruaphis noxia (Kurdjumov), Diruaphis tritici (Gillette), Diruaphis frequens (Walker), Diruaphis mexicana (McVicar Baker), Sipha flava (Forbes), Sipha elegans del Guercio, and Sitobion avenae (F.). In addition to the above-mentioned monoecious species, the common cereal aphids Rhopalosiphum padi (L.) and Rhopalosiphum maidis (Fitch) were included and successfully differentiated. DNA barcoding is a reliable alternative to traditional morphology in the identification of cereal aphids and their various life stages and morphs, including eggs. The application of DNA barcoding to aphid eggs found on grasses will be able to confirm whether D. noxia is now reproducing sexually in the United States after 20 yr of asexual reproduction.