Quantifying ELT-2 Protein Levels in Embryos

ELT-2 protein levels in embryos were measured as described previously (Van Fürden et al. 2004 (link); Wiesenfahrt et al. 2016 (link)), using the anti-ELT-2 monoclonal antibody 455-2A4. Images were recorded with a Hamamatsu Orca ER camera attached to a Zeiss Axioplan 2i microscope (40X objective); images for comparing different strains were collected with constant microscope/camera parameters. Total fluorescent intensity in the intestinal primordium (2E or 8E cell stage embryo) was measured (in arbitrary units) using ImageJ and corrected for background intensity measured elsewhere in the embryo. The probability that ELT-2 levels measured in one set of embryos have the same distribution of intensities as measured in JM247 embryos (single copy end-1p::elt-2 transgene) was calculated by the Wilcoxon-Mann-Whitney rank test (Marx et al. 2016 (link)) implemented online (https://ccb-compute2.cs.uni-saarland.de/wtest/).

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Wiesenfahrt T., Duanmu J., Snider F., Moerman D., Au V., Li-Leger E., Flibotte S., Parker D.M., Marshall C.J., Nishimura E.O., Mains P.E, & McGhee J.D. (2018). A Strategy To Isolate Modifiers of Caenorhabditis elegans Lethal Mutations: Investigating the Endoderm Specifying Ability of the Intestinal Differentiation GATA Factor ELT-2. G3: Genes|Genomes|Genetics, 8(5), 1425-1437.

Publication 2018

Orca ER camera

Anti antibody Cell Embryos Intestinal Microscope Orca Protein Strains Transgene

Corresponding Organization : University of Calgary

Other organizations : Canada's Michael Smith Genome Sciences Centre, University of British Columbia, Colorado State University

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Variable analysis

independent variables

Genotypes or strains of C. elegans embryos

dependent variables

ELT-2 protein levels in the intestinal primordium of C. elegans embryos

control variables

Microscope and camera parameters, including the objective (40X) and Hamamatsu Orca ER camera
Background intensity measured elsewhere in the embryo

controls

Positive control: JM247 embryos (single copy end-1p::elt-2 transgene)
Negative control: Not explicitly mentioned

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