title: Whole-mount in situ detection of microRNAs on Arabidopsis tissues using Zip Nucleic Acid probes.
abstract: MicroRNAs (miRNAs) affect fundamental processes of development. In plants miRNAs regulate organ development, transition to flowering, and responses to abiotic/biotic stresses. To understand the biological role of miRNAs, in addition to identifying their targeted transcripts, it is necessary to characterize the spatiotemporal regulation of their expression. Many methods have been used to define the set of organ-specific miRNAs by tissue dissection and miRNA profiling but none of them can describe their tissue and cellular distribution at the high resolution provided by in situ hybridization (ISH). This article describes the setup and optimization of a whole-mount ISH protocol to target endogenous miRNAs on intact Arabidopsis seedlings using DIG-labeled Zip Nucleic Acid (ZNA) oligonucleotide probes. Automation of the main steps of the procedure by robotized liquid handling has also been implemented in the protocol for best reproducibility of results, enabling running of ISH experiments at high throughput.
authors.affiliation: Department of Agronomy, Food, Natural Resources, Animals and Environment, University of Padova, 35020 Legnaro (Padova), Italy. maura.begheldo@unipd.it
authors.forename: M
authors.initials: M
authors.lastname: Begheldo
authors.forename: F A
authors.initials: FA
authors.lastname: Ditengou
authors.forename: G
authors.initials: G
authors.lastname: Cimoli
authors.forename: S
authors.initials: S
authors.lastname: Trevisan
authors.forename: S
authors.initials: S
authors.lastname: Quaggiotti
authors.forename: A
authors.initials: A
authors.lastname: Nonis
authors.forename: K
authors.initials: K
authors.lastname: Palme
authors.forename: B
authors.initials: B
authors.lastname: Ruperti
mesh_terms: D017360:Arabidopsis / Q000235:genetics* / Q000254:growth & development; D001331:Automation; D017403:In Situ Hybridization; D035683:MicroRNAs / Q000032:analysis*; D015345:Oligonucleotide Probes / Q000378:metabolism*; D036226:Seedlings / Q000235:genetics
chemical_list: D035683:MicroRNAs; D015345:Oligonucleotide Probes