Identification of Programmed Cell Death in situ in Individual Plant Cells in vivo Using a Chromosome Preparation Technique

Shun‐Bin Ning; Ling Wang; Yun‐Chun Song; Shunbin Ning

doi:10.1093/JEXBOT/53.369.651

Identification of Programmed Cell Death in situ in Individual Plant Cells in vivo Using a Chromosome Preparation Technique

Shun‐Bin Ning, Ling Wang, Yun‐Chun Song, Shunbin Ning

Wuhan University

Research output: Contribution to journal › Article › peer-review

Abstract

<div class="line" id="line-5"> <span style="font-family: Calibri, sans-serif;"> A simple procedure, which combines a chromosome preparation technique with an in situlabelling technique modified from fluorescence in situ hybridization (FISH), has been developed for in situ detection of plant programmed cell death (PCD) at the single&hyphen;cell level. After exposure of chromosomes and nuclei on slides by enzymolysis, Klenow or TdT was used to incorporate Bio&hyphen;dUTP or fluorescein&hyphen;dUTP at sites of DNA breaks. After Klenow&hyphen;mediated labelling, the signals were amplified by a cascade of antigen–antibody reaction according to the detection system of FISH. This method enables in situ detection of plant PCD in vivo morphologically and biochemically at the chromosome, nuclear and DNA levels without cell culture and histological sectioning. This technique permits labelling of DNA breaks with high sensitivity due to increased chromosome and nucleus exposure to the labelling solutions, as well as due to the immunological amplification of the signals. Moreover, the changes in the cells were easier to be observed because the spatial obstacle of the cell wall and its autofluorescence were eliminated. It is potentially useful for in situdetection of PCD in plant root meristematic cells triggered by various environmental abiotic factors. It is proposed that the root tip is a versatile in vivo system for studying PCD induced by environmental abiotic factors. </span></div>

Original language	American English
Journal	Journal of Experimental Botany
Volume	53
DOIs	https://doi.org/10.1093/JEXBOT/53.369.651
State	Published - Apr 1 2002

Keywords

Bio‐dUTP
Chromosome preparation
FISH
FITC
ISEL
PCD
PI
TUNEL
TdT
TdT‐mediated dUTP nick end‐labelling
biotin‐conjugated deoxyuridine triphosphate
fluorescein isothiocyanate.
fluorescence in situ hybridization
in situ end labelling
plants
programmed cell death
programmed cell death.
propidium iodide
terminal deoxynucleotidyl transferase

Disciplines

Biology
Botany

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@article{3c8c58e4c2844556a47987960699239e,

title = "Identification of Programmed Cell Death in situ in Individual Plant Cells in vivo Using a Chromosome Preparation Technique",

abstract = " A simple procedure, which combines a chromosome preparation technique with an\ in situlabelling technique modified from fluorescence\ in situ\ hybridization (FISH), has been developed for\ in situ\ detection of plant programmed cell death (PCD) at the single\&hyphen;cell level. After exposure of chromosomes and nuclei on slides by enzymolysis, Klenow or TdT was used to incorporate Bio\&hyphen;dUTP or fluorescein\&hyphen;dUTP at sites of DNA breaks. After Klenow\&hyphen;mediated labelling, the signals were amplified by a cascade of antigen\–antibody reaction according to the detection system of FISH. This method enables\ in situ\ detection of plant PCD\ in vivo\ morphologically and biochemically at the chromosome, nuclear and DNA levels without cell culture and histological sectioning. This technique permits labelling of DNA breaks with high sensitivity due to increased chromosome and nucleus exposure to the labelling solutions, as well as due to the immunological amplification of the signals. Moreover, the changes in the cells were easier to be observed because the spatial obstacle of the cell wall and its autofluorescence were eliminated. It is potentially useful for\ in situdetection of PCD in plant root meristematic cells triggered by various environmental abiotic factors. It is proposed that the root tip is a versatile\ in vivo\ system for studying PCD induced by environmental abiotic factors. ",

keywords = "Bio‐dUTP, Chromosome preparation, FISH, FITC, ISEL, PCD, PI, TUNEL, TdT, TdT‐mediated dUTP nick end‐labelling, biotin‐conjugated deoxyuridine triphosphate, fluorescein isothiocyanate., fluorescence in situ hybridization, in situ end labelling, plants, programmed cell death, programmed cell death., propidium iodide, terminal deoxynucleotidyl transferase",

author = "Shun‐Bin Ning and Ling Wang and Yun‐Chun Song and Shunbin Ning",

note = "A simple procedure, which combines a chromosome preparation technique with an in situ labelling technique modified from fluorescence in situ hybridization (FISH), has been developed for in situ detection of plant programmed cell death (PCD) at the single‐cell level.",

year = "2002",

month = apr,

day = "1",

doi = "10.1093/JEXBOT/53.369.651",

language = "American English",

volume = "53",

journal = "Journal of Experimental Botany",

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TY - JOUR

T1 - Identification of Programmed Cell Death in situ in Individual Plant Cells in vivo Using a Chromosome Preparation Technique

AU - Ning, Shun‐Bin

AU - Wang, Ling

AU - Song, Yun‐Chun

AU - Ning, Shunbin

N1 - A simple procedure, which combines a chromosome preparation technique with an in situ labelling technique modified from fluorescence in situ hybridization (FISH), has been developed for in situ detection of plant programmed cell death (PCD) at the single‐cell level.

PY - 2002/4/1

Y1 - 2002/4/1

N2 - A simple procedure, which combines a chromosome preparation technique with an in situlabelling technique modified from fluorescence in situ hybridization (FISH), has been developed for in situ detection of plant programmed cell death (PCD) at the single&hyphen;cell level. After exposure of chromosomes and nuclei on slides by enzymolysis, Klenow or TdT was used to incorporate Bio&hyphen;dUTP or fluorescein&hyphen;dUTP at sites of DNA breaks. After Klenow&hyphen;mediated labelling, the signals were amplified by a cascade of antigen–antibody reaction according to the detection system of FISH. This method enables in situ detection of plant PCD in vivo morphologically and biochemically at the chromosome, nuclear and DNA levels without cell culture and histological sectioning. This technique permits labelling of DNA breaks with high sensitivity due to increased chromosome and nucleus exposure to the labelling solutions, as well as due to the immunological amplification of the signals. Moreover, the changes in the cells were easier to be observed because the spatial obstacle of the cell wall and its autofluorescence were eliminated. It is potentially useful for in situdetection of PCD in plant root meristematic cells triggered by various environmental abiotic factors. It is proposed that the root tip is a versatile in vivo system for studying PCD induced by environmental abiotic factors.

AB - A simple procedure, which combines a chromosome preparation technique with an in situlabelling technique modified from fluorescence in situ hybridization (FISH), has been developed for in situ detection of plant programmed cell death (PCD) at the single&hyphen;cell level. After exposure of chromosomes and nuclei on slides by enzymolysis, Klenow or TdT was used to incorporate Bio&hyphen;dUTP or fluorescein&hyphen;dUTP at sites of DNA breaks. After Klenow&hyphen;mediated labelling, the signals were amplified by a cascade of antigen–antibody reaction according to the detection system of FISH. This method enables in situ detection of plant PCD in vivo morphologically and biochemically at the chromosome, nuclear and DNA levels without cell culture and histological sectioning. This technique permits labelling of DNA breaks with high sensitivity due to increased chromosome and nucleus exposure to the labelling solutions, as well as due to the immunological amplification of the signals. Moreover, the changes in the cells were easier to be observed because the spatial obstacle of the cell wall and its autofluorescence were eliminated. It is potentially useful for in situdetection of PCD in plant root meristematic cells triggered by various environmental abiotic factors. It is proposed that the root tip is a versatile in vivo system for studying PCD induced by environmental abiotic factors.

KW - Bio‐dUTP

KW - Chromosome preparation

KW - FISH

KW - FITC

KW - ISEL

KW - PCD

KW - PI

KW - TUNEL

KW - TdT

KW - TdT‐mediated dUTP nick end‐labelling

KW - biotin‐conjugated deoxyuridine triphosphate

KW - fluorescein isothiocyanate.

KW - fluorescence in situ hybridization

KW - in situ end labelling

KW - plants

KW - programmed cell death

KW - programmed cell death.

KW - propidium iodide

KW - terminal deoxynucleotidyl transferase

UR - https://doi.org/10.1093/jexbot/53.369.651

U2 - 10.1093/JEXBOT/53.369.651

DO - 10.1093/JEXBOT/53.369.651

M3 - Article

VL - 53

JO - Journal of Experimental Botany

JF - Journal of Experimental Botany

ER -

Identification of Programmed Cell Death in situ in Individual Plant Cells in vivo Using a Chromosome Preparation Technique

Abstract

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