Comparing the efficiency of six clearing methods in developing seeds of Arabidopsis thaliana

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00159816%3A_____%2F22%3A00077659" target="_blank" >RIV/00159816:_____/22:00077659 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216224:14740/22:00127169

Výsledek na webu

<a href="https://link.springer.com/article/10.1007/s00497-022-00453-4" target="_blank" >https://link.springer.com/article/10.1007/s00497-022-00453-4</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1007/s00497-022-00453-4" target="_blank" >10.1007/s00497-022-00453-4</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Comparing the efficiency of six clearing methods in developing seeds of Arabidopsis thaliana

Popis výsledku v původním jazyce

Key message ClearSee alpha and FAST9 were optimized for imaging Arabidopsis seeds up to the torpedo stages. The methods preserve the fluorescence of reporter proteins and seed shape, allowing phenotyping embryos in intact seeds. Tissue clearing methods eliminate the need for sectioning, thereby helping better understand the 3D organization of tissues and organs. In the past fifteen years, clearing methods have been developed to preserve endogenous fluorescent protein tags. Some of these methods (ClearSee, TDE, PEA-Clarity, etc.) were adapted to clear various plant species, with the focus on roots, leaves, shoot apical meristems, and floral parts. However, these methods have not been used in developing seeds beyond the early globular stage. Tissue clearing is problematic in post-globular seeds due to various apoplastic barriers and secondary metabolites. In this study, we compared six methods for their efficiency in clearing Arabidopsis thaliana seeds at post-globular embryonic stages. Three methods (TDE, ClearSee, and ClearSee alpha) have already been reported in plants, whereas the others (fsDISCO, FAST9, and CHAPS clear) are used in this context for the first time. These methods were assessed for seed morphological changes, clearing capacity, removal of tannins, and spectral properties. We tested each method in seeds from globular to mature stages. The pros and cons of each method are listed herein. ClearSee alpha appears to be the method of choice as it preserves seed morphology and prevents tannin oxidation. However, FAST9 with 60% iohexol as a mounting medium is faster, clears better, and appears suitable for embryonic shape imaging. Our results may guide plant researchers to choose a suitable method for imaging fluorescent protein-labeled embryos in intact Arabidopsis seeds.

Název v anglickém jazyce

Comparing the efficiency of six clearing methods in developing seeds of Arabidopsis thaliana

Popis výsledku anglicky

Key message ClearSee alpha and FAST9 were optimized for imaging Arabidopsis seeds up to the torpedo stages. The methods preserve the fluorescence of reporter proteins and seed shape, allowing phenotyping embryos in intact seeds. Tissue clearing methods eliminate the need for sectioning, thereby helping better understand the 3D organization of tissues and organs. In the past fifteen years, clearing methods have been developed to preserve endogenous fluorescent protein tags. Some of these methods (ClearSee, TDE, PEA-Clarity, etc.) were adapted to clear various plant species, with the focus on roots, leaves, shoot apical meristems, and floral parts. However, these methods have not been used in developing seeds beyond the early globular stage. Tissue clearing is problematic in post-globular seeds due to various apoplastic barriers and secondary metabolites. In this study, we compared six methods for their efficiency in clearing Arabidopsis thaliana seeds at post-globular embryonic stages. Three methods (TDE, ClearSee, and ClearSee alpha) have already been reported in plants, whereas the others (fsDISCO, FAST9, and CHAPS clear) are used in this context for the first time. These methods were assessed for seed morphological changes, clearing capacity, removal of tannins, and spectral properties. We tested each method in seeds from globular to mature stages. The pros and cons of each method are listed herein. ClearSee alpha appears to be the method of choice as it preserves seed morphology and prevents tannin oxidation. However, FAST9 with 60% iohexol as a mounting medium is faster, clears better, and appears suitable for embryonic shape imaging. Our results may guide plant researchers to choose a suitable method for imaging fluorescent protein-labeled embryos in intact Arabidopsis seeds.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10611 - Plant sciences, botany

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

N - Vyzkumna aktivita podporovana z neverejnych zdroju

Rok uplatnění

2022

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Plant Reproduction

ISSN

2194-7953

e-ISSN

2194-7961

Svazek periodika

35

Číslo periodika v rámci svazku

4

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

15

Strana od-do

279-293

Kód UT WoS článku

000884195100001

EID výsledku v databázi Scopus

—