Review: Plant Binary Vectors of Ti Plasmid in Agrobacterium tumefaciens with a Broad Host-Range Replicon of pRK2, pRi, pSa or pVS1

Abstract

This review\rchronicles the development of the plant binary vectors of Ti plasmid in Agrobacterium tumefaciens during the last\r30 years. A binary vector strategy was designed in 1983 to separate the T-DNA\rregion in a small plasmid from the virulence genes in avirulent T-DNA-less Ti plasmid. The small plant vectors with the\rT-DNA region have been simply now called binary Ti vectors. A binary Ti vector\rconsist of a broad host-range replicon for propagation in A. tumeraciens, an antibiotic resistance gene for bacterial\rselection and the T-DNA region that would be transferred to the plant genome\rvia the bacterial virulence machinery.\rThe T-DNA region delimited by the right and left border sequences contains an\rantibiotic resistance gene for plant selection, reporter gene, and/or any genes\rof interest. The ColEI replicon was also added to the plasmid backbone to\renhance the propagation in Escherichia\rcoli. A general trend in the binary vector development has been to increase the\rplasmid stability during a long co-cultivation period of A. tumefaciens with the target host plant tissues. A second trend\ris to understand the molecular mechanism of broad host-range replication, and\rto use it to reduce the size of plasmid for ease in cloning and for higher\rplasmid yield in E. coli. The broad\rhost-range replicon of VS1 was shown to be a choice of replicon over those of\rpRK2, pRi and pSA because of the superior stability and of small\rwell-defined replicon. Newly developed plant binary vectors pLSU has the small\rsize of plasmid backbone (4566 bp) consisting of VS1 replicon (2654 bp), ColE1\rreplicon (715 bp), a bacterial kanamycin (999 bp) or tetracycline resistance\rgene, and the T-DNA region (152 bp).