Combining laser-assisted microdissection (LAM) and RNA-seq allows to perform a comprehensive transcriptomic analysis of epidermal cells of Arabidopsis embryo
2018
Sakai, Kaori | Taconnat, Ludivine | Borrega, Néro | Yansouni, Jennifer | Brunaud, Véronique | Paysant-Le Roux, Christine | Delannoy, Etienne | Martin Magniette, Marie-Laure | Lepiniec, Loic | Faure, Jean Denis | Balzergue, Sandrine | Dubreucq, Bertrand | Institut Jean-Pierre Bourgin (IJPB) ; Institut National de la Recherche Agronomique (INRA)-AgroParisTech | Université Paris Saclay (COmUE) | Institut des Sciences des Plantes de Paris-Saclay (IPS2 (UMR_9213 / UMR_1403)) ; Institut National de la Recherche Agronomique (INRA)-Université Paris-Sud - Paris 11 (UP11)-Université Paris Diderot - Paris 7 (UPD7)-Université d'Évry-Val-d'Essonne (UEVE)-Centre National de la Recherche Scientifique (CNRS) | Mathématiques et Informatique Appliquées (MIA-Paris) ; Institut National de la Recherche Agronomique (INRA)-AgroParisTech | French State grant (LabEx Saclay Plant Sciences-SPS) [ANR-10-LABX-0040-SPS], managed by the French National Research Agency under an "Investments for the Future" program [ANR-11-IDEX-0003-02] | ANR-10-LABX-0040,SPS,Saclay Plant Sciences(2010) | ANR-11-IDEX-0003,IPS,Idex Paris-Saclay(2011)
International audience
Mostrar más [+] Menos [-]Inglés. Background: Genome- wide characterization of tissue- or cell-specific gene expression is a recurrent bottleneck in biology. We have developed a sensitive approach based on ultra-low RNA sequencing coupled to laser assisted microdissection for analyzing different tissues of the small Arabidopsis embryo. Methods and results: We first characterized the number of genes detected according to the quantity of tissue yield and total RNA extracted. Our results revealed that as low as 0.02 mm(2) of tissue and 50 pg of total RNA can be used without compromising the number of genes detected. The optimised protocol was used to compare the epidermal versus mesophyll cell transcriptomes of cotyledons at the torpedo- shaped stage of embryo development. The approach was validated by the recovery of well- known epidermal genes such <em>AtML1</em> or <em>AtPDF2</em> and genes involved in flavonoid and cuticular waxes pathways. Moreover, the interest and sensitivity of this approach were highlighted by the characterization of several transcription factors preferentially expressed in epidermal cells. Conclusion: This technical advance unlocks some current limitations of transcriptomic analyses and allows to investigate further and efficiently new biological questions for which only a very small amounts of cells need to be isolated. For instance, it paves the way to increasing the spatial accuracy of regulatory networks in developing small embryo of Arabidopsis or other plant tissues.
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