Fig. 2: Validation and benchmark of snRandom-seq using a human-mouse mixture sample. | Nature Communications

Fig. 2: Validation and benchmark of snRandom-seq using a human-mouse mixture sample.

From: High-throughput single nucleus total RNA sequencing of formalin-fixed paraffin-embedded tissues by snRandom-seq

Fig. 2

a Microfluidic encapsulation device for barcoding of nuclei. b Image of encapsulated droplet containing one bead, one nuclei, and reagents mix. c Electropherogram of 293T (human) and 3T3 (mouse) nuclei mixture cDNA library for Qsep100™ DNA Fragment Analyzer. Lower (20 bp) and upper (1 kb) markers were shown. d Barcode plot for identification of the barcodes that represent true nuclei (red line). Barcodes of the 293T-3T3 mixed nuclei were ordered from the largest to smallest gene counts. e Species-mixing scatter plot showing the single-nuclei capture efficiency and doublet rate of snRandom-seq. f Species specificity of UMIs in 293T-3T3 mixture. Identified 293T nuclei: n = 1157, identified 3T3 nuclei: n = 1086. Median of species specificity of UMIs in 293T was 0.992. Median of species specificity of UMIs in 3T3 was 0.986. g Percents of reads mapped to introns and exons. Violin plots and box plots showed the number of genes (h) and UMIs (i) detected in each 293T and 3T3 nucleus. Filtered 293T nuclei: n = 1085, filtered 3T3 nuclei: n = 1066. Data in the box plot corresponded to the first and third quartiles (lower and upper hinges) and median (center). j Saturation analysis of three methods. snRandom-seq used 293T and 3T3 nuclei; 10X Chromium Single Cell 3’ Solution V3 used 293 T and 3T3 cells; VASA-seq used 293T cells. k Read coverage along the gene body by the three methods. snRandom-seq used 293T nuclei; 10X Chromium Single Cell 3’ Solution V3 used 293T cells; VASA-seq used 293T cells. Data in (f, h, i) were presented as median values. Data in the box plot in (f, h, i) corresponded to the first (lower hinges) quartiles, third quartiles (upper hinges), and median (center). The upper whisker extended from the hinge to the maxima no further than 1.5 * IQR (interquartile range) from the hinge. The lower whisker extended from the hinge to the minima at most 1.5 * IQR of the hinge. IQR is the distance between the first and third quartiles. Source data are provided as a Source Data file.

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