Fluorescence Imaging of the Cytoskeleton in Plant Roots.
During the past two decades the use of live cytoskeletal probes has increased dramatically due to the introduction of the green fluorescent protein. However, to make full use of these live cell reporters it is necessary to implement simple methods to maintain plant specimens in optimal growing conditions during imaging. To image the cytoskeleton in living Arabidopsis roots, we rely on a system involving coverslips coated with nutrient supplemented agar where the seeds are directly germinated. This coverslip system can be conveniently transferred to the stage of a confocal microscope with minimal disturbance to the growth of the seedling. For roots with a larger diameter such as Medicago truncatula, seeds are first germinated in moist paper, grown vertically in between plastic trays, and roots mounted on glass slides for confocal imaging. Parallel with our live cell imaging approaches, we routinely process fixed plant material via indirect immunofluorescence. For these methods we typically use non-embedded vibratome-sectioned and whole mount permeabilized root tissue. The clearly defined developmental regions of the root provide us with an elegant system to further understand the cytoskeletal basis of plant development.
McDermott Center for Human Growth & Development, University of Texas Southwestern Medical Center, Dallas, TX, 75390, USA.
Julia
J
Dyachok
Plant Biology Division, The Samuel Roberts Noble Foundation, Inc., 2510 Sam Noble Parkway, Ardmore, OK, 73401, USA.
Ana
A
Paez-Garcia
Plant Biology Division, The Samuel Roberts Noble Foundation, Inc., 2510 Sam Noble Parkway, Ardmore, OK, 73401, USA.
Cheol-Min
CM
Yoo
Department of Biotechnology, Kalasalingam University, Krishnankoil, Tamil Nadu, 626126, India.
Karuppaiah
K
Palanichelvam
Plant Biology Division, The Samuel Roberts Noble Foundation, Inc., 2510 Sam Noble Parkway, Ardmore, OK, 73401, USA. eblancaflor@noble.org.
Elison B
EB
Blancaflor
D017360:Arabidopsis / Q000166:cytology*; D002470:Cell Survival; D003599:Cytoskeleton / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000378:metabolism; D007150:Immunohistochemistry; D046913:Medicago truncatula / Q000166:cytology*; D061848:Optical Imaging / Q000379:methods*; D018517:Plant Roots / Q000166:cytology*
D005456:Fluorescent Dyes
Actin; Arabidopsis; Fixed plant material; Green fluorescent protein; Immunofluorescence; Living cells; Medicago truncatula; Microtubules; Roots; Sectioning