3D fluorescent in situ hybridization using Arabidopsis leaf cryosections and isolated nuclei.
BACKGROUND
Fluorescent hybridization techniques are widely used to study the functional organization of different compartments within the mammalian nucleus. However, few examples of such studies are known in the plant kingdom. Indeed, preservation of nuclei 3D structure, which is required for nuclear organization studies, is difficult to fulfill.


RESULTS
We report a rapid protocol for fluorescent in situ hybridization (FISH) performed on 3D isolated nuclei and thin cryosectioned leaves of Arabidopsis thaliana. The use of direct labeling minimized treatment steps, shortening the overall procedure. Using image analysis, we measured different parameters related to nucleus morphology and overall 3D structure.


CONCLUSIONS
Our work describes a 3D-FISH protocol that preserves the 3D structure of Arabidopsis interphase nuclei. Moreover, we report for the first time FISH using cryosections of Arabidopsis leaves. This protocol is a valuable tool to investigate nuclear architecture and chromatin organization.
Tirichine
Leïla
L
Laboratoire de Biologie Cellulaire, INRA UR 501, IJPB, Route de Saint-Cyr, F-78026 Versailles, France. Valerie.Gaudin@versailles.inra.fr.
Andrey
Philippe
P
Biot
Eric
E
Maurin
Yves
Y
Gaudin
Valérie
V