5-4.4.3.2. Binary vector system

A plasmid carrying T-DNA containing LB and RB, called mini-Ti or micro-Ti can be sub- cloned in a small E. coli plasmid for ease of manipulation (Figure 5-4.4.3.2(b).).


The T-DNA of mini-Ti can be introduced into an Agrobacterium strain carrying a Ti plasmid from which the T-DNA has been removed but contains vir region (Figure 5-4.4.3.2(a).). The vir genes function in trans, causing transfer of the recombinant T-DNA to the plant genome.


The T-DNA plasmid can be introduced into Agrobacterium by triparental mating or by a more simple transformation procedure, such as electroporation.

Figure 5-4.4.3.2: A binary vector system (a) A plasmid containing vir region but no T-DNA, therefore no T-DNA transfer takes place in plant genome. (b) Another plasmid containing T-DNA with Right border (RB) and Left border (LB) but no vir genes. Vir function is supplied in trans by former plasmid.

In Ti-plasmid transformation system, the T-DNA is maintained on a shuttle vector with a broad host range origin of replication, such as RK2 (which functions in both A. tumefaciens and E. coli), or separate origin for each species.

An independently replicating vector is advantageous because maintenance of the T-DNA does not rely on recombination. The copy number of binary vector is not determined by the Ti plasmid, making the identification of transformants much easier.