Transient gene expression in plant cells mediated by Agrobacterium tumefaciens: Application for the analysis of virulence loci

Yasushi Yoshioka, Yoshito Takahashi, Ken Matsuoka, Kenzo Nakamura, Jun Koizumi, Mineo Kojima, Yasunori Machida

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

A simple system is described for detection of the transfer of T-DNA from Agrobacterium cells to suspension-cultured tobacco BY-2 cells. A modified reporter gene for β-glucuronidase (GUS) that contained an intron sequence was introduced into the T-DNA region such that the GUS protein could be synthesized in plant cells only after transfer of the T-DNA to plant nuclei. When BY-2 cells were co-cultured with Agrobacterium cells that contained the modified reporter gene, transient synthesis of GUS protein was observed between 36 and 48 h after the onset of co-culture. The level of GUS activity reached a plateau within as little as 48 h. This temporal profile of GUS activation suggests that the transient activity might have been due to expression of the GUS gene in the T-DNA that had been transferred to the plant nuclei but had not yet been integrated into the plant chromosomes. Levels of transient GUS activity were also examined with various vir mutants of Agrobacterium and in a mutant with an altered chromosomal acvB gene, the gene for a protein that has been postulated to function outside bacterial cells. During co-culture with virB, virD2, virD4 and acvB mutants, GUS activity remained at background levels, and the GUS activity in the case of the virE2 mutant was thirty-fold lower than with the wild type. On the basis of these results, we discuss the roles of these genes during infection by Agrobacterium of plant cells.

Original languageEnglish
Pages (from-to)782-789
Number of pages8
JournalPlant and Cell Physiology
Volume37
Issue number6
DOIs
Publication statusPublished - Sep 1996

Fingerprint

Agrobacterium tumefaciens
Glucuronidase
Plant Cells
Agrobacterium radiobacter
Virulence
virulence
Gene Expression
gene expression
loci
Agrobacterium
mutants
cells
DNA
coculture
reporter genes
Coculture Techniques
genes
Reporter Genes
Plant Chromosomes
proteins

All Science Journal Classification (ASJC) codes

  • Physiology
  • Plant Science
  • Cell Biology

Cite this

Transient gene expression in plant cells mediated by Agrobacterium tumefaciens : Application for the analysis of virulence loci. / Yoshioka, Yasushi; Takahashi, Yoshito; Matsuoka, Ken; Nakamura, Kenzo; Koizumi, Jun; Kojima, Mineo; Machida, Yasunori.

In: Plant and Cell Physiology, Vol. 37, No. 6, 09.1996, p. 782-789.

Research output: Contribution to journalArticle

Yoshioka, Yasushi ; Takahashi, Yoshito ; Matsuoka, Ken ; Nakamura, Kenzo ; Koizumi, Jun ; Kojima, Mineo ; Machida, Yasunori. / Transient gene expression in plant cells mediated by Agrobacterium tumefaciens : Application for the analysis of virulence loci. In: Plant and Cell Physiology. 1996 ; Vol. 37, No. 6. pp. 782-789.
@article{a1b8328871b54bec86d0a3aea2f9a592,
title = "Transient gene expression in plant cells mediated by Agrobacterium tumefaciens: Application for the analysis of virulence loci",
abstract = "A simple system is described for detection of the transfer of T-DNA from Agrobacterium cells to suspension-cultured tobacco BY-2 cells. A modified reporter gene for β-glucuronidase (GUS) that contained an intron sequence was introduced into the T-DNA region such that the GUS protein could be synthesized in plant cells only after transfer of the T-DNA to plant nuclei. When BY-2 cells were co-cultured with Agrobacterium cells that contained the modified reporter gene, transient synthesis of GUS protein was observed between 36 and 48 h after the onset of co-culture. The level of GUS activity reached a plateau within as little as 48 h. This temporal profile of GUS activation suggests that the transient activity might have been due to expression of the GUS gene in the T-DNA that had been transferred to the plant nuclei but had not yet been integrated into the plant chromosomes. Levels of transient GUS activity were also examined with various vir mutants of Agrobacterium and in a mutant with an altered chromosomal acvB gene, the gene for a protein that has been postulated to function outside bacterial cells. During co-culture with virB, virD2, virD4 and acvB mutants, GUS activity remained at background levels, and the GUS activity in the case of the virE2 mutant was thirty-fold lower than with the wild type. On the basis of these results, we discuss the roles of these genes during infection by Agrobacterium of plant cells.",
author = "Yasushi Yoshioka and Yoshito Takahashi and Ken Matsuoka and Kenzo Nakamura and Jun Koizumi and Mineo Kojima and Yasunori Machida",
year = "1996",
month = "9",
doi = "10.1093/oxfordjournals.pcp.a029013",
language = "English",
volume = "37",
pages = "782--789",
journal = "Plant and Cell Physiology",
issn = "0032-0781",
publisher = "Oxford University Press",
number = "6",

}

TY - JOUR

T1 - Transient gene expression in plant cells mediated by Agrobacterium tumefaciens

T2 - Application for the analysis of virulence loci

AU - Yoshioka, Yasushi

AU - Takahashi, Yoshito

AU - Matsuoka, Ken

AU - Nakamura, Kenzo

AU - Koizumi, Jun

AU - Kojima, Mineo

AU - Machida, Yasunori

PY - 1996/9

Y1 - 1996/9

N2 - A simple system is described for detection of the transfer of T-DNA from Agrobacterium cells to suspension-cultured tobacco BY-2 cells. A modified reporter gene for β-glucuronidase (GUS) that contained an intron sequence was introduced into the T-DNA region such that the GUS protein could be synthesized in plant cells only after transfer of the T-DNA to plant nuclei. When BY-2 cells were co-cultured with Agrobacterium cells that contained the modified reporter gene, transient synthesis of GUS protein was observed between 36 and 48 h after the onset of co-culture. The level of GUS activity reached a plateau within as little as 48 h. This temporal profile of GUS activation suggests that the transient activity might have been due to expression of the GUS gene in the T-DNA that had been transferred to the plant nuclei but had not yet been integrated into the plant chromosomes. Levels of transient GUS activity were also examined with various vir mutants of Agrobacterium and in a mutant with an altered chromosomal acvB gene, the gene for a protein that has been postulated to function outside bacterial cells. During co-culture with virB, virD2, virD4 and acvB mutants, GUS activity remained at background levels, and the GUS activity in the case of the virE2 mutant was thirty-fold lower than with the wild type. On the basis of these results, we discuss the roles of these genes during infection by Agrobacterium of plant cells.

AB - A simple system is described for detection of the transfer of T-DNA from Agrobacterium cells to suspension-cultured tobacco BY-2 cells. A modified reporter gene for β-glucuronidase (GUS) that contained an intron sequence was introduced into the T-DNA region such that the GUS protein could be synthesized in plant cells only after transfer of the T-DNA to plant nuclei. When BY-2 cells were co-cultured with Agrobacterium cells that contained the modified reporter gene, transient synthesis of GUS protein was observed between 36 and 48 h after the onset of co-culture. The level of GUS activity reached a plateau within as little as 48 h. This temporal profile of GUS activation suggests that the transient activity might have been due to expression of the GUS gene in the T-DNA that had been transferred to the plant nuclei but had not yet been integrated into the plant chromosomes. Levels of transient GUS activity were also examined with various vir mutants of Agrobacterium and in a mutant with an altered chromosomal acvB gene, the gene for a protein that has been postulated to function outside bacterial cells. During co-culture with virB, virD2, virD4 and acvB mutants, GUS activity remained at background levels, and the GUS activity in the case of the virE2 mutant was thirty-fold lower than with the wild type. On the basis of these results, we discuss the roles of these genes during infection by Agrobacterium of plant cells.

UR - http://www.scopus.com/inward/record.url?scp=0029808780&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0029808780&partnerID=8YFLogxK

U2 - 10.1093/oxfordjournals.pcp.a029013

DO - 10.1093/oxfordjournals.pcp.a029013

M3 - Article

AN - SCOPUS:0029808780

VL - 37

SP - 782

EP - 789

JO - Plant and Cell Physiology

JF - Plant and Cell Physiology

SN - 0032-0781

IS - 6

ER -