Myogenic conversion and transcriptional profiling of embryonic blastomeres in Caenorhabditis elegans.
Myogenesis has proven to be a powerful paradigm for understanding cell fate specification and differentiation in many model organisms. This includes the nematode Caenorhabditis elegans for which the genetic, cellular, and molecular tools have allowed an in-depth understanding of muscle development. One tool not yet available in C. elegans is a robust, pure and prolific cell culture system to study myogenesis. As an alternative, this chapter describes a method by which the cell fates of early, uncommitted blastomeres in the embryo are converted to a myogenic lineage. This technique permits the nearly synchronous induction of myogenesis in vivo with the potential to generate a nearly homogeneous population of cells. Coupled with the RNA isolation and cDNA amplification methods that are also described, one can now profile gene expression throughout myogenesis using any platform of choice (e.g. expression arrays, next generation sequencing). Although limited by the artificial nature of this developing mass of muscle inside the eggshell, blastomere conversion and transcriptional profiling is a very powerful tool to investigate changes in gene expression associated with myogenesis in C. elegans that is applicable to many different cell types. When coupled with next generation sequencing, the method has the potential to yield a very high-resolution map of changes in gene expression throughout myogenesis.
Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892, United States.
Tetsunari
T
Fukushige
Michael
M
Krause
D000818:Animals; D001757:Blastomeres / Q000378:metabolism; D017173:Caenorhabditis elegans / Q000196:embryology* / Q000235:genetics* / Q000378:metabolism; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D024510:Muscle Development; D009218:Myosins / Q000648:ultrastructure; D016133:Polymerase Chain Reaction; D013194:Staining and Labeling
D009218:Myosins