README_movie_filescontrol_EMBD0000_GLS_longcontrol_EMBD0000_MS_longaph-1_EMBD0022_GLS-1aph-1_EMBD0022_MSapl-1_EMBD0082_GLSapl-1_EMBD0082_MSaps-1_EMBD0013_GLSaps-1_EMBD0013_MSarx-1_EMBD0006_GLS-2arx-1_EMBD0006_MSarx-3_EMBD0055_GLS-2arx-3_EMBD0055_MSast-1_EMBD0074_GLSast-1_EMBD0074_MSceh-43_EMBD0018_GLSceh-43_EMBD0018_MSdie-1_EMBD0009_GLSdie-1_EMBD0009_MSdsh-2_EMBD0038_GLSdsh-2_EMBD0038_MSeff-1_EMBD0042_GLSeff-1_EMBD0042_MSelt-1_EMBD0016_GLSelt-1_EMBD0016_MSemb-9_EMBD0062_GLSemb-9_EMBD0062_MSgex-2_EMBD0005_GLSgex-2_EMBD0005_MSgrh-1_EMBD0012_GLSgrh-1_EMBD0012_MShch-1_EMBD0041_GLShch-1_EMBD0041_MShmg-3_EMBD0003_GLShmg-3_EMBD0003_MShmr-1_EMBD0057_GLShmr-1_EMBD0057_MSlag-1_EMBD0068_GLSlag-1_EMBD0068_MSlam-3_EMBD0014_GLSlam-3_EMBD0014_MSlet-19_EMBD0004_GLSlet-19_EMBD0004_MSlet-381_EMBD0015_GLSlet-381_EMBD0015_MSlet-4_EMBD0026_GLSlet-4_EMBD0026_MSlet-413_EMBD0054_GLSlet-413_EMBD0054_MSlet-502_EMBD0007_GLSlet-502_EMBD0007_MSlin-26_EMBD0052_GLSlin-26_EMBD0052_MSlit-1_EMBD0387_GLSlit-1_EMBD0387_MSmbk-2_EMBD0077_GLSmbk-2_EMBD0077_MSmes-1_EMBD0043_GLSmes-1_EMBD0043_MSmex-3_EMBD0063_GLSmex-3_EMBD0063_MSmom-2_EMBD0386_GLSmom-2_EMBD0386_MSpal-1_EMBD0053_GLSpal-1_EMBD0053_MSpha-1_EMBD0025_GLSpha-1_EMBD0025_MSpha-4_EMBD0002_GLSpha-4_EMBD0002_MSplrg-1_EMBD0076_GLSplrg-1_EMBD0076_MSpop-1_EMBD0388_GLSpop-1_EMBD0388_MSpxn-2_EMBD0039_GLSpxn-2_EMBD0039_MSsem-4_EMBD0080_GLSsem-4_EMBD0080_MSvab-1_EMBD0027_GLSvab-1_EMBD0027_MSvab-19_EMBD0029_GLSvab-19_EMBD0029_MSwrm-1_EMBD0385_GLSwrm-1_EMBD0385_MS

The C. elegans embryo is an important model for analyzing mechanisms of cell fate specification and tissue morphogenesis. Sophisticated lineaging approaches for analyzing embryogenesis have been developed but are labor-intensive and do not naturally integrate morphogenetic readouts. To enable the rapid classification of developmental phenotypes, we developed a high-content method that employs two custom strains: a Germ Layer strain expressing nuclear markers in the ectoderm, mesoderm and endoderm/pharynx, and a Morphogenesis strain expressing markers labeling epidermal cell junctions and the neuronal cell surface. We describe a procedure that allows simultaneous live imaging of development in 80-100 embryos and provide a custom program that generates cropped, oriented image stacks of individual embryos to facilitate analysis. We demonstrate the utility of our method by perturbing 40 previously characterized developmental genes in variants of the two strains containing RNAi-sensitizing mutations. The resulting datasets yielded distinct, reproducible signature phenotypes for a broad spectrum of genes involved in cell fate specification and morphogenesis. Our analysis additionally provides new in vivo evidence for MBK-2 function in mesoderm fate specification and LET-381 function in elongation.