Cell-to-cell variability does not appear to explain the low abundance of the lncRNAs in our survey. (a) Distribution of RNA single molecules counts (bins, columns; Red intensity: fractional density of molecule counts across the population) for the 64 lncRNA-cell pairs in the validated set that are quantitative (rows, Methods). Cell type color coding: orange - HeLa, blue - hFF, purple - hLF. Left bins are sized 3 (0 to 50 molecules ), where right in bins are sized 10 (50 to 300 molecules). A heterogeneously expressed lncRNA (XLOC_003526) and a homogenously expressed lncRNA (XLOC_006922), are pointed by black arrows and referenced in figures b and c. (b, c) The relationship between the mean molecule count (X axis) vs. median molecule count (Y axis, b) or vs. variability in molecule counts (Y axis, coefficient of variation, c) for the 64 lncRNA-cell pairs in the quantitative validated set (red), mRNA set 1 (green circles; Methods) and mRNA set 2 (green diamonds; Methods). A linear regression line in b (black) supports the consistency of the majority of transcript-cell pairs with a unimodal distribution (Y = 0.87X-1.25, Pearson r = 0.96). Dotted line is Y = X. Black curve in (c) is the theoretic Poisson distribution. Four transcripts marked (1 to 4) are analyzed further in d and e. LncRNA pairs with mean >170 (less than 10% of all pairs) are not presented, but show a similar pattern on a log scale. (d) Fluorescence micrographs of single molecule RNA FISH of a homogenously expressed lncRNA (1-XLOC_006922; top left) and mRNA (2-FOXF1; top right) and of a heterogeneously expressed lncRNA (3- XLOC_003526; bottom left) and mRNA (4 - CCNA2; bottom right). XLOC_003526 and CCNA2 are both heterogeneous but do not correlate with each other based on co-staining in two colors. Scale bar, 5 μm. (e) Molecule count distributions for each of the example transcript 1 to 4.