ASSOCIATION OF SINGLE-STRANDED TRANSFERRED DNA FROM AGROBACTERIUM-TUMEFACIENS WITH TOBACCO CELLS

During the inception of crown gall tumorigenesis, the transferred DNA (T-DNA) is processed from the Ti (tumor inducing) plasmid of Agrobacte rium tumefaciens and is transferred to plant cells. T-DNA processing a nd transfer require the induction of vir (virulence) genes by phenolic compounds secreted by wounded plant cells. After vir gene induction, both single-stranded (T-strands) and double-stranded forms of processe d T-DNA accumulate in the bacteria. Although current models favor the transfer of T-strands to plants, there has yet been no experimental ev idence to show this. In this paper, we show that T-strands disappear f rom acetosyringone-induced A. tumefaciens within 30 min of bacterial c ocultivation with tobacco protoplasts. PCR analysis of T-DNA associate d with protoplasts indicates that single-stranded, but not double-stra nded, T-DNA can be detected in the plant cells within 30 min of bacter ial cocultivation. Control experiments show that this T-DNA does not o riginate from lysed contaminating bacterial cells. T-DNA transfer depe nds on a functional bacterial virB operon. Protoplast infections using an A. tumefaciens virE mutant result in a low level of accumulation o f T-strands in the plant cells.