Bleckmann, Andrea and Dresselhaus, Thomas (2016) Fluorescent whole-mount RNA in situ hybridization (F-WISH) in plant germ cells and the fertilized ovule. METHODS, 98. pp. 66-73. ISSN 1046-2023, 1095-9130
Full text not available from this repository. (Request a copy)Abstract
First evidence on gene function and regulation is provided by the cellular expression pattern in complex tissues. However, to understand the activity of a specific gene, it is essential to analyze the regulatory network, which controls the spatio-temporal translation pattern during the entire life span of the transcribed mRNA. To explore mechanisms which control mRNA abundance and localization in space and time, it is necessary to visualize mRNAs quantitatively with a subcellular resolution, without sectioning the tissues. We have adapted and optimized a protocol for colorimetric whole-mount RNA in situ hybridization (WISH) using egg cell-specific digoxigenin (DIG) labeled probes (Hejatko et al., 2006) [1] on ovules and early seeds of Arabidopsis. Furthermore, we established a fluorescent whole-mount RNA in situ hybridization (F-WISH) protocol, which allows mRNA visualization on a subcellular level. The polar localized mRNA of SBT4.13, encoding a subtilase, was identified using this protocol. Both methods are described and discussed in detail. Additionally a (F)-WISH flow-chart is provided along with a troubleshooting table. (C) 2015 Elsevier Inc. All rights reserved.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | MESSENGER; LOCALIZATION; COMPLEXES; mRNA; WISH; TSA System; F-WISH; Subtilase; Ovule |
| Subjects: | 500 Science > 570 Life sciences 500 Science > 580 Botanical sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Pflanzenwissenschaften > Lehrstuhl für Zellbiologie und Pflanzenphysiologie |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 01 Mar 2019 14:12 |
| Last Modified: | 25 Mar 2019 13:27 |
| URI: | https://pred.uni-regensburg.de/id/eprint/3171 |
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