The lineaging of fluorescently-labeled Caenorhabditis elegans embryos with StarryNite and AceTree.
Lineage analysis of Caenorhabditis elegans is a powerful tool for characterizing developmental phenotypes and embryonic gene-expression patterns. We present a detailed protocol for the lineaging of embryos by computational analysis of 4D images of embryos that ubiquitously express histone-GFP (green fluorescent protein) fusion proteins through the 350 cell stage followed by manual editing. We describe how to optimize imaging settings for this purpose, the use of the lineage-extraction software, StarryNite, and the lineage-editing software, AceTree. In addition, we describe a useful polymer bead mounting technique for C. elegans embryos that has several advantages compared with the standard agar pad mounting technique. The protocol requires about 1 h of user time spread over 2 days to generate the raw lineage, and an additional 2 or 4 h to edit the lineage to the 194- or 350-cell stage, respectively.
Murray
John Isaac
JI
Department of Genome Sciences, University of Washington Box 357730, Seattle, Washington 98195, USA.
Bao
Zhirong
Z
Boyle
Thomas J
TJ
Waterston
Robert H
RH
D000818:Animals; D017173:Caenorhabditis elegans / Q000196:embryology*; D019070:Cell Lineage / Q000502:physiology*; D004625:Embryo, Nonmammalian / Q000196:embryology / Q000378:metabolism; D020869:Gene Expression Profiling / Q000379:methods*; D049452:Green Fluorescent Proteins; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D010641:Phenotype; D012984:Software
D049452:Green Fluorescent Proteins