studying of sequence similarity between different Ti plasmids in 1981. Between 1980–2000, the characterization of the T-DNA region and the 'vir' region was also pursued. Studies into the T-DNA region determined their process of transfer and identified genes allowing the synthesis of plant hormones and opines. Separately, early work aimed to determine the functions of the genes encoded in the 'vir' region - these were broadly categorized into those that allowed bacterial-host interactions and those that enabled T-DNA delivery. Replication, partitioning and maintenance The replication, partitioning and maintenance of the Ti plasmid depends on the repABC gene cassette, which is mainly made up of three genes: repA, repB and repC. repA and repB each encode for proteins involved in plasmid partitioning, while repC encodes a replication initiator. These genes are expressed from 4 different promoters located upstream of repA. repE encodes for a small antisense RNA and is located between repB and repC. Additionally, there is a partitioning site (parS) and an origin of replication (oriV) present within the repABC cassette. Replication of the Ti plasmid The replication of the Ti plasmid is driven by the RepC initiator protein (), which possesses two protein domains: an N-terminal domain (NTD) that binds to DNA and a C-terminal domain (CTD). Mutational analyses have shown that without a functional RepC protein, the Ti plasmid is unable to replicate. Meanwhile, the oriV sequence is around 150 nucleotides in length and is found within the repC gene. Laboratory experiments have shown that the RepC protein binds to this region, suggesting its role as the origin of replication. Therefore, while the complete process behind the replication of the Ti plasmid has not been fully described, the initial step of replication would likely depend on the expression of RepC and its binding to oriV. Of note, the RepC protein only acts in cis, where it only drives the replication of the plasmid it is encoded in and not any other plasmid also present in the bacterial cell. Partitioning of the Ti plasmid The partitioning system of the Ti plasmid is similar to the ParA/ParB system used in other plasmids and bacterial chromosomes and is thought to act in the same way. Mutations in either of the proteins RepA or RepB have resulted in a decrease in plasmid stability, indicating their role and importance in plasmid partitioning. The ability of RepA to form filaments allows it to create