Aarhus University Seal / Aarhus Universitets segl

Optimizing protocols for arabidopsis shoot and root protoplast cultivation

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

Standard

Optimizing protocols for arabidopsis shoot and root protoplast cultivation. / Pasternak, Taras; Paponov, Ivan A.; Kondratenko, Serhii.

In: Plants, Vol. 10, No. 2, 375, 02.2021, p. 1-17.

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaperJournal articleResearchpeer-review

Harvard

APA

CBE

MLA

Vancouver

Author

Pasternak, Taras ; Paponov, Ivan A. ; Kondratenko, Serhii. / Optimizing protocols for arabidopsis shoot and root protoplast cultivation. In: Plants. 2021 ; Vol. 10, No. 2. pp. 1-17.

Bibtex

@article{7102af33017b4b4fae37778871cf2044,
title = "Optimizing protocols for arabidopsis shoot and root protoplast cultivation",
abstract = "Procedures for the direct regeneration of entire plants from a shoot and root protoplasts of Arabidopsis thaliana have been optimized. The culture media for protoplast donor-plant cultivation and protoplast culture have been adjusted for optimal plant growth, plating efficiency, and promotion of shoot regeneration. Protocols have been established for the detection of all three steps in plant regeneration: (i) chromatin relaxation and activation of auxin biosynthesis, (ii) cell cycle progression, and (iii) conversion of cell-cycle active cells to totipotent ones. The competence for cell division was detected by DNA replication events and required high cell density and high concentrations of the auxinic compound 2,4-D. Cell cycle activity and globular structure formation, with subsequent shoot induction, were detected microscopically and by labeling with fluorescent dye Rhodamine123. The qPCR results demonstrated significantly upregulated expression of the genes responsible for nuclear reorganization, auxin responses, and auxin biosynthesis during the early stage of cell reprogramming. We further optimized cell reprogramming with this protocol by applying glutathione (GSH), which increases the sensitivity of isolated mesophyll protoplasts to cell cycle activation by auxin. The developed protocol allows us to investigate the molecular mechanism of the de-differentiation of somatic plant cells.",
keywords = "Arabidopsis thaliana, Auxin, Protoplasts, Reprograming",
author = "Taras Pasternak and Paponov, {Ivan A.} and Serhii Kondratenko",
note = "Funding Information: Funding: This research was supported by the National Academy of Agrarian Sciences of Ukraine in the course of the second level assignment (type of assignment 18.00.01.01 F) of scientific research program No. 18 {"}Vegetable and melon growing{"}. This work was supported by Bundesministerium f{\"u}r Bildung und Forschung (BMBF 349 Microsystems). Publisher Copyright: {\textcopyright} 2021 by the authors. Licensee MDPI, Basel, Switzerland. Copyright: Copyright 2021 Elsevier B.V., All rights reserved.",
year = "2021",
month = feb,
doi = "10.3390/plants10020375",
language = "English",
volume = "10",
pages = "1--17",
journal = "Plants",
issn = "2223-7747",
publisher = "MDPI AG",
number = "2",

}

RIS

TY - JOUR

T1 - Optimizing protocols for arabidopsis shoot and root protoplast cultivation

AU - Pasternak, Taras

AU - Paponov, Ivan A.

AU - Kondratenko, Serhii

N1 - Funding Information: Funding: This research was supported by the National Academy of Agrarian Sciences of Ukraine in the course of the second level assignment (type of assignment 18.00.01.01 F) of scientific research program No. 18 "Vegetable and melon growing". This work was supported by Bundesministerium für Bildung und Forschung (BMBF 349 Microsystems). Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. Copyright: Copyright 2021 Elsevier B.V., All rights reserved.

PY - 2021/2

Y1 - 2021/2

N2 - Procedures for the direct regeneration of entire plants from a shoot and root protoplasts of Arabidopsis thaliana have been optimized. The culture media for protoplast donor-plant cultivation and protoplast culture have been adjusted for optimal plant growth, plating efficiency, and promotion of shoot regeneration. Protocols have been established for the detection of all three steps in plant regeneration: (i) chromatin relaxation and activation of auxin biosynthesis, (ii) cell cycle progression, and (iii) conversion of cell-cycle active cells to totipotent ones. The competence for cell division was detected by DNA replication events and required high cell density and high concentrations of the auxinic compound 2,4-D. Cell cycle activity and globular structure formation, with subsequent shoot induction, were detected microscopically and by labeling with fluorescent dye Rhodamine123. The qPCR results demonstrated significantly upregulated expression of the genes responsible for nuclear reorganization, auxin responses, and auxin biosynthesis during the early stage of cell reprogramming. We further optimized cell reprogramming with this protocol by applying glutathione (GSH), which increases the sensitivity of isolated mesophyll protoplasts to cell cycle activation by auxin. The developed protocol allows us to investigate the molecular mechanism of the de-differentiation of somatic plant cells.

AB - Procedures for the direct regeneration of entire plants from a shoot and root protoplasts of Arabidopsis thaliana have been optimized. The culture media for protoplast donor-plant cultivation and protoplast culture have been adjusted for optimal plant growth, plating efficiency, and promotion of shoot regeneration. Protocols have been established for the detection of all three steps in plant regeneration: (i) chromatin relaxation and activation of auxin biosynthesis, (ii) cell cycle progression, and (iii) conversion of cell-cycle active cells to totipotent ones. The competence for cell division was detected by DNA replication events and required high cell density and high concentrations of the auxinic compound 2,4-D. Cell cycle activity and globular structure formation, with subsequent shoot induction, were detected microscopically and by labeling with fluorescent dye Rhodamine123. The qPCR results demonstrated significantly upregulated expression of the genes responsible for nuclear reorganization, auxin responses, and auxin biosynthesis during the early stage of cell reprogramming. We further optimized cell reprogramming with this protocol by applying glutathione (GSH), which increases the sensitivity of isolated mesophyll protoplasts to cell cycle activation by auxin. The developed protocol allows us to investigate the molecular mechanism of the de-differentiation of somatic plant cells.

KW - Arabidopsis thaliana

KW - Auxin

KW - Protoplasts

KW - Reprograming

UR - http://www.scopus.com/inward/record.url?scp=85100768916&partnerID=8YFLogxK

U2 - 10.3390/plants10020375

DO - 10.3390/plants10020375

M3 - Journal article

C2 - 33672063

AN - SCOPUS:85100768916

VL - 10

SP - 1

EP - 17

JO - Plants

JF - Plants

SN - 2223-7747

IS - 2

M1 - 375

ER -