Accumulation of numerous cellular T-DNA sequences in the genus Diospyros by multiple rounds of natural transformation

Horizontal gene transfer (HGT) is an important phenomenon in the evolutionary history of plants. Natural transformation by Agrobacterium is a special case of HGT and leads to the insertion of cellular T-DNA (cT-DNA) sequences, for example, in Diospyros lotus. The genus Diospyros contains about 795 species with economically important members, like different types of persimmon (D. kaki, D. lotus, and D. virginiana) and ebony (e.g., D. ebenum). Whole genome sequences (WGS) from D. kaki, D. oleifera, D. lotus, and D. virginiana were investigated for cT-DNAs. These four species belong to one clade and contain 15 different cT-DNAs (DiTA to DiTO). The hexaploid species D. kaki cv. “Xiaoguo-tianshi” contains seven types of cT-DNA (DiTA to DiTG) on 27 of 42 homeologs, adding up to 628 kb of cT-DNA. Five of these seven cT-DNAs are non-fixed, as shown by empty chromosomal insertion sites. The evolutionary history of the Diospyros cT-DNAs was reconstructed using the divergence of their inverted repeats. Insert age varied from 3 to 12 million years. Partial cT-DNA sequences were detected in 35 additional species from five Diospyros clades. Our data highlight the unexpectedly large scale of natural Agrobacterium transformation in Diospyros and demonstrate the necessity of whole genome approaches for studies on the origin and evolution of cT-DNAs.