title: Association of single-stranded transferred DNA from Agrobacterium tumefaciens with tobacco cells.
abstract: 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 processing 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 protoplasts. 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.
authors.affiliation: Department of Biological Sciences, Purdue University, West Lafayette, IN 47907.
authors.forename: V M
authors.initials: VM
authors.lastname: Yusibov
authors.forename: T R
authors.initials: TR
authors.lastname: Steck
authors.forename: V
authors.initials: V
authors.lastname: Gupta
authors.forename: S B
authors.initials: SB
authors.lastname: Gelvin
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics*; D001483:Base Sequence; D002478:Cells, Cultured; D017931:DNA Primers / Q000737:chemistry; D004277:DNA, Single-Stranded / Q000378:metabolism*; D018014:Gene Transfer Techniques; D005798:Genes, Bacterial; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D010947:Plants, Toxic; D010957:Plasmids; D014026:Nicotiana
chemical_list: D017931:DNA Primers; D004277:DNA, Single-Stranded