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Association of single-stranded transferred DNA from Agrobacterium tumefaciens with tobacco cells

Yusibov, V.M., Steck, T.R., Gupta, V., Gelvin, S.B.
Proceedings of the National Academy of Sciences of the United States of America 1994 v.91 no.8 pp. 2994-2998
protoplasts, structural genes, Agrobacterium radiobacter, plasmids, acetosyringone, Nicotiana tabacum, virulence, single-stranded DNA
During the inception of crown gall tumorigenesis, the transferred DNA (T-DNA) is processed from the Ti (tumor inducing) plasmid of Agrobacterium tumefaciens and is transferred to plant cells. T-DNA p and 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 processed T-DNA accumulate in the bacteria. Although current models favor the transfer of T-strands to plants, there has yet been no experimental evidence to show this. In this paper, we show that T-strands disappear from acetosyringone-induced A. tumefaciens within 30 min of bacterial cocultivation with tobacco protoplast. PCR analysis of T-DNA associated with protoplasts indicates that single-stranded, but not double-stranded, T-DNA can be detected in the plant cells within 30 min of bacterial cocultivation. Control experiments show that this T-DNA does not originate from lysed contaminating bacterial cells. T-DNA transfer depends on a functional bacterial virB operon. Protoplast infections using an A. tumefaciens virE mutant result in a low level of accumulation of T-strands in the plant cells.