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Combining an original method for preserving RNA expression in situ with an effective RNA extraction method makes it possible to study gene expression in any banana fruit tissue

Published online by Cambridge University Press:  09 June 2009

Ludivine Lassois ,
Luc de Lapeyre de Bellaire  and
Haïssam Jijakli
Ludivine Lassois
Affiliation:
Gembloux Agric. Univ., Plant Pathol. Unit, Passage des Déportés 2, B-5030 Gembloux, Belgium
Luc de Lapeyre de Bellaire
Affiliation:
CIRAD, Persyst, UPR Syst. Banan. Ananas, TA B-26 / PS4, Blvd. de la Lironde, 34398 Montpellier Cedex 5, France
Haïssam Jijakli
Affiliation:
Gembloux Agric. Univ., Plant Pathol. Unit, Passage des Déportés 2, B-5030 Gembloux, Belgium
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Abstract

Introduction. RNA isolation is a prerequisite to studying gene expression in banana and to understanding changes occurring in response to the environment. Standard extraction methods do not efficiently extract RNA from plants such as banana, with high levels of phenolics, carbohydrates, or other compounds that bind to and/or coprecipitate with RNA. Materials and methods. Five to seven RNA extraction methods were compared. Four crown-tissue storage methods were also compared. cDNA-AFLP was used to ensure that the obtained RNA was of sufficient quality for molecular applications and that RNA expression was unaltered by in situ storage. Results and discussion. The modified hot-borate method proved to be the best RNA extraction method, allowing high yields of good quality, undegraded RNA from the crown, fruit peel and pulp at all stages of ripening. The RNA obtained by this method was of sufficient quality for molecular applications such as cDNA-AFLP that give highly reproducible results. Freeze-drying of fresh tissues and tissue conservation in hot-borate buffer, two original storage methods, appear appropriate for preserving RNA in situ without ultra-low temperature. The RNA obtained was of high quality, undegraded, and useful for all downstream applications. The genome expression profile obtained by cDNA-AFLP analysis was unaltered by these methods for storing collected tissues. Conclusion. By applying all the suggested procedures in this work, it is possible to store and study gene expression in any banana fruit tissue, whatever the maturity stage, without affecting the RNA expression level.

Keywords

BelgiumMusa sp.bananasfreeze-dryingRNAstorageextractionBelgiqueMusa sp.bananelyophilisationARNstockageextraction
Type
Research Article
Information
Fruits , Volume 64 , Issue 3 , May 2009 , pp. 127 - 137
Copyright
© CIRAD, EDP Sciences, 2009

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