A variety of plant species found in nature contain agrobacterial T-DNAs in their genomes which they transmit in a series of sexual generations. Such T-DNAs are called cellular T-DNAs (cT-DNAs). cT-DNAs have been discovered in dozens of plant genera, and are suggested to be used in phylogenetic studies, since they are well-defined and unrelated to other plant sequences. Their integration into a particular chromosomal site indicates a founder event and a clear start of a new clade. cT-DNA inserts do not disseminate in the genome after insertion. They can be large and old enough to generate a range of variants, thereby allowing the construction of detailed trees. Unusual cT-DNAs (containing the rolB/C-like gene) were found in our previous study in the genome data of two Vaccinium L. species. Here, we present a deeper study of these sequences in Vaccinium L. Molecular-genetic and bioinformatics methods were applied for sequencing, assembly, and analysis of the rolB/C-like gene. The rolB/C-like gene was discovered in 26 new Vaccinium species and Agapetes serpens (Wight) Sleumer. Most samples were found to contain full-size genes. It allowed us to develop approaches for the phasing of cT-DNA alleles and reconstruct a Vaccinium phylogenetic relationship. Intra- and interspecific polymorphism found in cT-DNA makes it possible to use it for phylogenetic and phylogeographic studies of the Vaccinium genus.