Simplified method for cell-specific gene expression analysis in Caenorhabditis elegans

In the neural circuit functional identities of individual neurons are mainly specified by their differential gene expression patterns. Unveiling functional roles of each neuron requires cell-specific interrogation of neural circuitry in the context of gene expressions. The mRNA tagging strategy in Caenorhabditis elegans is a powerful technique, in which cell-specific transcripts can be isolated by co-immunoprecipitating the complexes of mRNAs and epitope-tagged poly(A) binding protein (3 x FLAG-PAB-1), expressed in target neurons. However, the conventional protocol requires laborious and time-consuming procedures; chromosomal integration of gene encoding 3 x FLAG-PAB-1 and bleaching of obtained integrant animals for the isolation of huge amounts of synchronized animals. In this paper, we have presented a simplified methodology for cell-specific mRNA tagging analysis in C elegans. We show that mRNA tagging was achieved using transgenic animals expressing 3 x FLAG-PAB-1 as an extrachromosomal array under the control of the flp-18 promoter, without the chromosomal integration procedure. Furthermore, we successfully isolated cell-specific mRNAs from adult transgenic animals synchronously grown from eggs laid by gravid adults during a time window of 3 h. This simplification facilitates the implementation of cell-specific gene expression analysis of C. elegans, which contributes to the understanding of neural circuitry at a cell-specific resolution. (C) 2014 Elsevier Inc. All rights reserved.