Extended Data Fig. 9: Cell loss and barcode dropouts.
From: Clonal relations in the mouse brain revealed by single-cell and spatial transcriptomics
a, Workflow for counting the total number of barcoded EGFP+ cells in tissue sections. We counted EGFP+ cells separately in cortex (CX, red), hippocampus (HC, green) and striatum (STR, blue) in 50 µm thick coronal sections. For each region, we determined the labeling density (EGFP+ cells/mm3) and obtained the total number of cells by multiplying labeling density with volume estimations of the respective region. b, The average number of EGFP+ cells per brain is 566,000 ± 29,500 cells (n = 9 sections). Graph represents average number of cells per brain, error bars represent SD, colored dots represent individual data points corresponding to the number of labelled cells per section and brain. c, Recovery rate of labelled cells and barcodes at each step of the TREX protocol. Upon tissue dissociation isolated all EGFP+ cells contained in the suspension using bulk sorting (orange bar, ‘FACS’) and sorted cells were recovered via centrifugation. All recovered cells were transferred into the wells of a 10X Chromium chip (purple bar, ‘Loaded’) for droplet encapsulation. Following quality control and filtering, a subset of cells was contained in the final dataset (magenta bar, ‘Dataset’). A large fraction of cells had a barcode (lightgreen bar, ‘cloneID+’) of which many were contained in clones defined as a group of at least two cells sharing the same cloneID (yellow bar, ‘Clones’). d, Average recovery rate of cells and barcodes for all brains compared to the average total number of barcoded cells present in all three analyzed forebrain regions. e, Recovery rate of barcodes at each step of the Space-TREX protocol. Based on widefield images recorded for each section before library preparation, we could determine the total number of EGFP+ cells in a section (darkgreen bar, ‘EGFP cells’) and the number of those cells located in spots containing capture probes (orange bar, ‘EGFP cells in spots’). We displayed the final number of spots containing a cloneID (purple bar “Spots with cloneID”). f, Average recovery rate of cells and barcodes for all sections.
