Agrobacterium tumefaciens (Ti plasmid) — Virulent strains carry large tumor-inducing (Ti) plasmids that mediate DNA transfer to plant cells. How many distinct border sequence sets required for gene transfer are present on a classical Ti plasmid (i.e., the left and right T-DNA borders)?

Introduction / Context:
Agrobacterium tumefaciens transfers a defined DNA segment, called transfer DNA (T-DNA), from its Ti plasmid into the nuclear genome of wounded plant cells. This process depends on short, specific border sequences that mark the left and right ends of the T-DNA to be mobilized. Understanding the number and role of these borders is foundational in plant genetic engineering and binary vector design.

Given Data / Assumptions:

• A Ti plasmid harbors a T-DNA region destined for transfer.
• Transfer requires precise recognition sites (border repeats) by the Vir protein machinery.
• Only DNA between the borders is transferred and integrated.

Concept / Approach:
The essential cis-acting signals for T-DNA excision and transfer are two short, largely imperfect direct repeats known as the left border (LB) and right border (RB). VirD1/VirD2 endonuclease complex nicks these borders to generate the transferable single-stranded T-strand. Therefore, there are two sets of required border sequences per T-DNA: one at each end.

Step-by-Step Solution:

Verification / Alternative check:
Binary vector systems retain LB and RB around cargo genes; removing either border abolishes transfer, confirming the requirement for two borders flanking the insert.

Why Other Options Are Wrong:

Common Pitfalls:
Confusing vir gene locations (in trans) with cis borders; vir genes are not transferred, while the bordered segment is.

Final Answer:
Two sets of sequences necessary for gene transfer.