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<div>Rose rosette is one of the most economically important diseases of roses. It is caused by rose rosette virus (RRV), a hepta-segmented, negative-sense RNA virus, member of the genus <i>Emaravirus</i>; transmitted by the eriophyid mite <i>Phyllocoptes fructiphilus</i>. Little is known about the virus and studies on its population structure and evolution are crucial to better understand the epidemiology of virus and disease. We implemented a multiplex amplicon sequencing method to quantify polymorphisms in the RRV U.S. population. On hundred and seven (107) RRV isolates collected from 19 states were used in the study. After reverse transcription, samples were submitted to PCR using virus-specific primers developed to amplify the complete virus genome and sequenced on Illumina platforms. Bioinformatics analysis provided information of more than 95% of the genome length for more than 90 isolates. The mean genetic distance among isolates was the highest for RNA 5 and the lowest for RNAs 2 and 3. Phylogenetic trees constructed for all genomic RNAs. Isolates were grouped into distinct clusters with high bootstrap support with geography having minimum impact in virus evolution, possibly because of the movement of infected material across state lines or because the plant genotypes are the driving force shaping virus populations.</div>