LIVE CONFOCAL IMAGING OF GROWING ARABIDOPSIS ROOTS
2School of Mechanical Engineering, Tel Aviv University, Tel Aviv, Israel
Plant roots exhibit a number of complex behaviors, while performing the double task of providing mechanical anchorage to the plant and taking up nutrients (water and minerals) from the soil to be delivered to the whole plant. In doing this, the root grows in soil and explores the environment looking for areas which are richer in resources, while redirecting its growth to avoid obstacles like rocks. An important feature is that the growth happens only in a small region at the tip, the back of the root being anchored in the soil without moving anymore.
The plant we use in our studies is Arabidopsis thaliana, a tiny plant whose roots in the early stages of growth present a diameter on the order of 150µm; this size makes it suitable for observation via confocal microscopy. Our interest is twofold:
(1) we want to measure the mechanical forces exerted by the root by growing in the embedding medium, and compare them to those exerted by a mechanical probe of similar dimensions pushed in the same medium. To do this, we grow an Arabidopsis th. root in agarose gel embedding fluorescent microbeads, and observe its growth in a confocal microscope, measuring the displacement of the beads with Digital Volume Correlation (DVC) [1,2] technique.
(2) Another aspect we are interested in is the process of decision making in plant roots – how they respond to chemical stimuli in terms of growth path. We can observe this process “on the flight” by using a microfluidic device [3] where the root grows inside a microchannel while being exposed to an asymmetric medium flow (left and right side of the root sensing two different media).
Both of the projects present non trivial challenges due to the potentially adverse growing conditions for the plant, which needs more light and humidity than inside the confocal microscope. Designing and creating ad hoc set-ups helped us circumvent such difficulties and perform experiments of the duration of typically 12 hours.
[1] Franck C., Maskarinec S.A., Ravichandran C., Tirrell D.A., "Quantifying Cellular Traction Forces in Three Dimensions", PNAS, 106:22108 - 22113, 2009.
[2] Lesman A., Notbohm J., Tirrell D.A., Ravichandran G., "Contractile forces regulate cell division in three-dimensional environments", J Cell Biol (2014) 205 (2): 155–162..
[3] Grossmann G., Guo W.J., Ehrhardt D.W., Frommer W.B., Sit R.V., Quake S.R., Meier M., "The RootChip: an integrated microfluidic chip for plant science", The plant cell 23 (12), 4234-4240, 2011.