Long noncoding RNAs (lncRNAs) exhibit diverse functions, including regulation of development. Here the authors combined genome-wide mapping of SMAD3 occupancy with expression analysis to identify lncRNAs induced by activin signaling during endoderm differentiation of human embryonic stem cells (hESCs). They found that DIGIT is divergent to Goosecoid (GSC) and expressed during endoderm differentiation. Using single molecule RNA fluorescent in-situ hybridization (FISH) and SAO-based imaging with the StellarVision instrument, the authors found that DIGIT transcripts are primarily retained in the nucleus while GSC mRNA transcripts are most abundant in the cytoplasm. The large field of view enabled by SAO allowed quantification of this distribution across 50 cells, revealing that about 90% of DIGIT transcripts were retained in the nucleus compared to 15% of GSC transcripts. Nuclear localization of DIGIT provided further support that DIGIT is not an mRNA, and this analysis also revealed that DIGIT and GSC are induced in the same cells during differentiation.