AbstractAquifoliaceae is the largest family in the campanulid order Aquifoliales. It consists of a single genus,Ilex, the hollies, which is the largest woody dioecious genus in the angiosperms. Most species are in East Asia or South America. The taxonomy and evolutionary history remain unclear due to the lack of a robust species-level phylogeny. We produced the first complete chloroplast genomes in this family, including sevenIlexspecies, by Illumina sequencing of long-range PCR products and subsequent reference-guidedde novoassembly. These genomes have a typical bicyclic structure with a conserved genome arrangement and moderate divergence. The total length is 157,741 bp and there is one large single-copy region (LSC) with 87,109 bp, one small single-copy with 18,436 bp, and a pair of inverted repeat regions (IR) with 52,196 bp. A total of 144 genes were identified, including 96 protein-coding genes, 40 tRNA and 8 rRNA. Thirty-four repetitive sequences were identified inIlex pubescens, with lengths >14 bp and identity >90%, and 11 divergence hotspot regions that could be targeted for phylogenetic markers. This study will contribute to improved resolution of deep branches of theIlexphylogeny and facilitate identification ofIlexspecies.