AbstractVincetoxicum mongolicumMaxim. (1876), is a perennial medicinal herb, widely distributed in the Loess Plateau of China. Here, we sequenced, assembled, and annotated the complete chloroplast (cp) genome ofV. mongolicum, and compared the highly variable gene regions and phylogenetic positions betweenV. mongolicumand other related species. Results showed that the complete cp genome ofV. mongolicumwas 160,157 bp in length, containing a large single copy (LSC) region of 91,263 bp, a pair of inverted repeats (IR) region of 23,892 bp, and a small single copy (SSC) region of 21,110 bp. The GC content accounts for 37.8%, and we annotated 131 single genes, which include 86 protein-coding genes, 8 rRNA genes, and 37 tRNA genes. By comparing and analyzing the variable region of the cp gene ofV. mongolicumand otherVincetoxicum, we found that the variable sequences ofrpoC1-rpoB,ycf4-cemA,ndhF,ndhF-rpl32, andrpl32-ccsAfragments were highly significant, which could be targeted as the DNA barcodes for evidence ofV. mongolicumand its relatives in Apocynaceae. Maximum-likelihood (ML) phylogenetic tree analysis elucidated thatV. mongolicumwas sister toV. pycnostelmawith strong support. Our results provide useful information for future phylogenetic studies and plastid super-barcodes of the family Apocynaceae.