Fig. 4

Surface-FISHseq analysis of PBMCs and functional tests of maxRNAs in primary human monocytes. a The Surface-FISHseq experimental workflow for the 3 technical variations. b A Venn diagram of maxRNAs identified by each of the 3 technical variations of Surface-FISHseq: Surface-FISHseq-membrane (blue), Surface-FISHseq-FACS (green), and Surface-FISHseq-Psoralen (orange). c–f Read distributions of Surface-FISHseq test libraries (red tracks) and control libraries (blue tracks) on (c) FNDC3B and (e) CTSS. d, f Expanded views of the 3′UTR regions of the two genes. Probes track (bottom track): locations and IDs of antisense oligonucleotides for functional tests. g Effects of probe incubation on the average monocyte attachment levels (normalized to the no-probe control (black), y-axis) of dArt4 control probeset (dArt4), the randomized 20 nt control probeset (random 20-mer), and the antisense probe-sets against the 11 Surface-FISHseq identified targets (gray columns). Each probe-set is comprised of twenty-five 20 nt antisense oligonucleotide probes. Error bar: standard error of the mean. N: number of replicate experiments. Each condition was compared to no-probe control. **Bonferroni-adjusted p < 0.001, ***Bonferroni-adjusted p < 0.0001, ****Bonferroni-adjusted p < 0.00001. h, i Effects of individual antisense probes to the average monocyte attachment levels (y-axis) in no-probe control (black), dArt4 control (red), and by each FNDC3B probe (gray columns, indexed by E1-E4, U1-U5 corresponding to locations in d) and each CTSS probe (E1, E2, U1-U9 corresponding to the locations in panel f). Each condition was compared to no-probe control. **Bonferroni-adjusted p < 0.001, ***Bonferroni-adjusted p < 0.0001