Time-lapse Fluorescence Imaging of Arabidopsis Root Growth with Rapid Manipulation of The Root Environment Using The RootChip
- Stanford Univ., CA (United States). Carnegie Inst. for Science. Dept. of Plant Biology
- Howard Hughes Medical Inst., Chevy Chase, MD (United States); Stanford Univ., CA (United States). Dept. of Appleid Physics and Bioengineering; Univ. of Freiburg (Germany). Center for Biological Signaling Studies (BIOSS). Dept. of Microsystems Engineering (IMTEK)
- Howard Hughes Medical Inst., Chevy Chase, MD (United States); Stanford Univ., CA (United States). Dept. of Appleid Physics and Bioengineering
The root functions as the physical anchor of the plant and is the organ responsible for uptake of water and mineral nutrients such as nitrogen, phosphorus, sulfate and trace elements that plants acquire from the soil. If we want to develop sustainable approaches to producing high crop yield, we need to better understand how the root develops, takes up a wide spectrum of nutrients, and interacts with symbiotic and pathogenic organisms. To accomplish these goals, we need to be able to explore roots in microscopic detail over time periods ranging from minutes to days. We developed the RootChip, a polydimethylsiloxane (PDMS)- based microfluidic device, which allows us to grow and image roots from Arabidopsis seedlings while avoiding any physical stress to roots during preparation for imaging1 (Figure 1). The device contains a bifurcated channel structure featuring micromechanical valves to guide the fluid flow from solution inlets to each of the eight observation chambers2 . This perfusion system allows the root microenvironment to be controlled and modified with precision and speed. The volume of the chambers is approximately 400 nl, thus requiring only minimal amounts of test solution. Here we provide a detailed protocol for studying root biology on the RootChip using imaging-based approaches with real time resolution. Roots can be analyzed over several days using time lapse microscopy. Roots can be perfused with nutrient solutions or inhibitors, and up to eight seedlings can be analyzed in parallel. This system has the potential for a wide range of applications, including analysis of root growth in the presence or absence of chemicals, fluorescence-based analysis of gene expression, and the analysis of biosensors, e.g. FRET nanosensors3.
- Research Organization:
- Carnegie Inst. of Washington, Washington, DC (United States)
- Sponsoring Organization:
- USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division; National Science Foundation (NSF)
- Grant/Contract Number:
- FG02-04ER15542; MCB 1021677
- OSTI ID:
- 1628632
- Journal Information:
- Journal of Visualized Experiments, Journal Issue: 65; ISSN 1940-087X
- Publisher:
- MyJoVE Corp.Copyright Statement
- Country of Publication:
- United States
- Language:
- English
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