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Binary Ti vectors are the plasmid vectors of choice in Agrobacterium-mediated plant transformation protocols. The pGreen series of binary Ti vectors are configured for ease-of-use and to meet the demands of a wide range of transformation procedures for many plant species. This plasmid system allows any arrangement of selectable marker and reporter gene at the right and left T-DNA borders without compromising the choice of restriction sites for cloning, since the pGreen cloning sites are based on the well-known pBluescript general vector plasmids. Its size and copy number in Escherichia coli offers increased efficiencies in routine in vitro recombination procedures. pGreen can replicate in Agrobacterium only if another plasmid, pSoup, is co-resident in the same strain. pSoup provides replication functions in trans for pGreen. The removal of RepA and Mob functions has enabled the size of pGreen to be kept to a minimum. Versions of pGreen have been used to transform several plant species with the same efficiencies as other binary Ti vectors. Information on the pGreen plasmid system is supplemented by an Internet site (http://www.pgreen.ac.uk) through which comprehensive information, protocols, order forms and lists of different pGreen marker gene permutations can be found.

Stem internodes of three diploid potato genotypes (lines HH260, C and E) were transformed with binary Agrobacterium strains, containing the A. rhizogenes wild type plasmid pRil855 together with the plasmid construct pBI 121, to introduce a set of selectable and reporter marker genes (coding for hairy root phenotype, hormone autotrophy, opine synthesis, kanamycin resistance and P-glucuronidase activity). Genetic marker lines were produced at the level of root clone, plant (regenerated from root clone segments) and cell suspension culture (established from callus induced on leaf segments). Transformation frequencies and the expression of transformation marker characters were dependent on the genotype, the physiological state of the internodal explants and the Agrobacterium strain. Root clones, derived from stem internodes producing transformed roots in high numbers, generally showed a complete set of marker characters and prolific growth, in contrast to the root clones that originated from less productive stem internodes. Shoot regeneration was achieved from the genotypes HH260 and C, but not from genotype E. Loss of one or more marker characters (opine synthesis, kanamycin resistance, GUS activity) was observed in half of the regenerants, as compared to their original root clones. Cell suspension cultures showed expression of all marker characters present in the original transformed plant. The majority of the transformed marker lines, at the level of root clone, regenerated plant, or cell suspension culture, had maintained the original diploid level.