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Carnegie Institution for Science

5 ARTICLES PUBLISHED IN JoVE

Bioengineering

Time-lapse Fluorescence Imaging of Arabidopsis Root Growth with Rapid Manipulation of The Root Environment Using The RootChip

Guido Grossmann¹, Matthias Meier^2,3,4, Heather N. Cartwright¹, Davide Sosso¹, Stephen R. Quake^2,3, David W. Ehrhardt¹, Wolf B. Frommer¹

¹Department of Plant Biology, Carnegie Institution for Science, ²Howard Hughes Medical Institute, ³Departments of Applied Physics and Bioengineering, Stanford University , ⁴Department of Microsystems Engineering (IMTEK) and Center for Biological Signaling Studies (BIOSS), University of Freiburg

This article provides a protocol for cultivation of Arabidopsis seedlings in the RootChip, a microfluidic imaging platform that combines automated control of growth conditions with microscopic root monitoring and FRET-based measurement of intracellular metabolite levels.

Biology

A New Approach for the Comparative Analysis of Multiprotein Complexes Based on ¹⁵N Metabolic Labeling and Quantitative Mass Spectrometry

Kerstin Trompelt¹, Janina Steinbeck¹, Mia Terashima^1,2, Michael Hippler¹

¹Institute of Plant Biology and Biotechnology, University of Münster, ²Department of Plant Biology, Carnegie Institution for Science

The described comparative, quantitative proteomic approach aims at obtaining insights into the composition of multiprotein complexes under different conditions and is demonstrated by comparing genetically different strains. For quantitative analysis equal volumes of different fractions from a sucrose density gradient are mixed and analyzed by mass spectrometry.

JoVE Journal

Isolation of Giant Lampbrush Chromosomes from Living Oocytes of Frogs and Salamanders

Joseph G. Gall¹, Zehra F. Nizami¹

¹Department of Embryology, Carnegie Institution for Science

We present simple techniques for isolating giant transcriptionally active lampbrush chromosomes from living oocytes of frogs and salamanders. We describe how to observe these chromosomes "alive" by phase contrast or differential interference contrast, and how to fix them for fluorescent in situ hybridization or immunofluorescent staining.

Biology

High-fat Feeding Paradigm for Larval Zebrafish: Feeding, Live Imaging, and Quantification of Food Intake

Jessica P. Otis¹, Steven A. Farber^1,2

¹Department of Embryology, Carnegie Institution for Science, ²Department of Biology, Johns Hopkins University

Zebrafish are emerging as a valuable model of dietary lipid processing and metabolic disease. Described are protocols of lipid-rich larval feeds, live imaging of dietary fluorescent lipid analogs, and quantification of food intake. These techniques can be applied to a variety of screening, imaging, and hypothesis driven inquiry techniques.

Biology

Fast Colony Forming Unit Counting in 96-Well Plate Format Applied to the Drosophila Microbiome

Ren Dodge¹, William B. Ludington^1,2

¹Department of Embryology, Carnegie Institution for Science, ²Department of Biology, Johns Hopkins University

This method quantifies microbial abundance using a 96-well plate format to plate colony forming units (CFUs) and is applied to the Drosophila microbiome in whole fly homogenate samples. CFUs are counted with an automated image analysis software provided here.