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Impact on the Sustainable Development Goals (SDGs)

Analysis of institutional authors

González-García MpCorresponding AuthorBustillo-Avendaño EAuthorSanchez-Corrionero AAuthorDel Pozo JcAuthorMoreno-Risueno MaAuthor

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April 29, 2020
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Fluorescence-activated cell sorting using the D-root device and optimization for scarce and/or non-accessible root cell populations

Publicated to: Plants-Basel. 9 (4): E499- - 2020-04-01 9(4), DOI: 10.3390/plants9040499

Authors:

Gonzalez-Garcia, Mary-Paz; Bustillo-Avendano, Estefano; Sanchez-Corrionero, Alvaro; del Pozo, Juan C; Moreno-Risueno, Miguel A
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Affiliations

Centre for Plant Biotechnology and Genomics UPM - INIA - Author
Univ Politecn Madrid, Ctr Biotecnol & Genom Plantas, Inst Nacl Invest & Tecnol Agr & Alimentaria, Campus Montegancedo, Madrid 28223, Spain - Author

Abstract

© 2020 by the authors. Licensee MDPI, Basel, Switzerland. Fluorescence-activated cell sorting (FACS) is a technique used to isolate specific cell populations based on characteristics detected by flow cytometry. FACS has been broadly used in transcriptomic analyses of individual cell types during development or under different environmental conditions. Different protoplast extraction protocols are available for plant roots; however, they were designed for accessible cell populations, which normally were grown in the presence of light, a non-natural and stressful environment for roots. Here, we report a protocol using FACS to isolate root protoplasts from Arabidopsis green fluorescent protein (GFP)-marked lines using the minimum number of enzymes necessary for an optimal yield, and with the root system grown in darkness in the D-Root device. This device mimics natural conditions as the shoot grows in the presence of light while the roots grow in darkness. In addition, we optimized this protocol for specific patterns of scarce cell types inside more differentiated tissues using the mCherry fluorescent protein. We provide detailed experimental protocols for effective protoplasting, subsequent purification through FACS, and RNA extraction. Using this RNA, we generated cDNA and sequencing libraries, proving that our methods can be used for genome-wide transcriptomic analyses of any cell-type from roots grown in darkness.
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Keywords

cell-typed-rootfacsfounder cellslateral rootsArabidopsisAuxinCell-typeCytokininD-rootDifferentiationExpressionFacsFlow-cytometryFounder cellsInitiationIntegrationLateral rootsLife on landLightProtoplastsRoot protoplasts

Quality index

Bibliometric impact. Analysis of the contribution and dissemination channel

The work has been published in the journal Plants-Basel due to its progression and the good impact it has achieved in recent years, according to the agency WoS (JCR), it has become a reference in its field. In the year of publication of the work, 2020, it was in position 47/235, thus managing to position itself as a Q1 (Primer Cuartil), in the category Plant Sciences.

Independientemente del impacto esperado determinado por el canal de difusión, es importante destacar el impacto real observado de la propia aportación.

Según las diferentes agencias de indexación, el número de citas acumuladas por esta publicación hasta la fecha 2026-04-27:

  • Google Scholar: 8
  • WoS: 6
  • Scopus: 9
  • Europe PMC: 7
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Impact and social visibility

From the perspective of influence or social adoption, and based on metrics associated with mentions and interactions provided by agencies specializing in calculating the so-called "Alternative or Social Metrics," we can highlight as of 2026-04-27:

  • The use, from an academic perspective evidenced by the Altmetric agency indicator referring to aggregations made by the personal bibliographic manager Mendeley, gives us a total of: 29.

With a more dissemination-oriented intent and targeting more general audiences, we can observe other more global scores such as:

  • The Total Score from Altmetric: 5.
  • The number of mentions on the social network X (formerly Twitter): 6 (Altmetric).

It is essential to present evidence supporting full alignment with institutional principles and guidelines on Open Science and the Conservation and Dissemination of Intellectual Heritage. A clear example of this is:

  • The work has been submitted to a journal whose editorial policy allows open Open Access publication.
  • Assignment of a Handle/URN as an identifier within the deposit in the Institutional Repository: https://oa.upm.es/93922/

As a result of the publication of the work in the institutional repository, statistical usage data has been obtained that reflects its impact. In terms of dissemination, we can state that, as of

  • Views: 34
  • Downloads: 10
Continuing with the social impact of the work, it is important to emphasize that, due to its content, it can be assigned to the area of interest of ODS 15 - Life on land, with a probability of 65% according to the mBERT algorithm developed by Aurora University.
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Leadership analysis of institutional authors

There is a significant leadership presence as some of the institution’s authors appear as the first or last signer, detailed as follows: First Author (GONZALEZ GARCIA, MARIA DE LA PAZ) and Last Author (MORENO RISUEÑO, MIGUEL ANGEL).

the author responsible for correspondence tasks has been GONZALEZ GARCIA, MARIA DE LA PAZ.

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Project objectives

Los objetivos perseguidos en esta aportación son analizar la aplicación de la técnica de clasificación celular por citometría de flujo (FACS) para aislar protoplastos de raíces de Arabidopsis cultivadas en condiciones naturales simuladas por el dispositivo D-Root; optimizar el protocolo de extracción de protoplastos utilizando el mínimo número de enzimas para maximizar el rendimiento; adaptar el método para la identificación y aislamiento de poblaciones celulares escasas o inaccesibles mediante marcadores fluorescentes como GFP y mCherry; caracterizar detalladamente los procedimientos experimentales para la purificación celular y extracción de ARN; y validar la utilidad del protocolo para generar bibliotecas de secuenciación y análisis transcriptómicos a nivel genómico de tipos celulares específicos en raíces cultivadas en oscuridad.
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Most relevant results

Este estudio presenta un protocolo optimizado para la clasificación celular por citometría de flujo (FACS) aplicado a raíces de Arabidopsis cultivadas en condiciones que simulan la oscuridad natural mediante el dispositivo D-Root. Los resultados más relevantes incluyen: 1) el uso del mínimo número de enzimas para obtener un rendimiento óptimo de protoplastos; 2) la adaptación del método para aislar poblaciones celulares escasas y específicas marcadas con proteína fluorescente mCherry; 3) la demostración de la viabilidad del protocolo para la extracción de ARN y generación de bibliotecas de secuenciación; y 4) la confirmación de que estas técnicas permiten análisis transcriptómicos a nivel genómico de células radiculares cultivadas en oscuridad, evitando el estrés inducido por la luz.
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