(a-e) Violin plots showing a comparison between manual CEL-Seq2 and the robotic version at different volume reductions for the distributions of (a) the number of transcripts per mESC, (b) the fraction of recovered ERCC1 spike-in RNA, (c) the number of genes per mESC, (d) Pearson’s correlation coefficient between measured and actual spike-in numbers, and (e) Pearson’s correlation coefficient between spike-in levels measured in distinct cells. In (a-e) violin plots represent the density distribution of the data. Overlaid boxplots show the median (white dot) and the interquartile range (box limits). The whiskers extend to the most extreme data point, within 1.5 times the interquartile range from the box. Outliers are indicated. The sample size in (a-e) 48 cells for each group from n=1 experiments. (f, g) Dependence of sequencing efficiency on sequence composition. A regression was calculated of the average sequenced spike-in number on the actual spike-in number, setting the intercept to zero. The scatter plots show the dependence of the deviation of the measured spike-in level from the regression line, normalized by the average expression, on (f) percentage GC content and (g) sequence length. Data points for 96 ERCC1 spike-in sequences are shown in (f) and (g). Shown data are from one experiment.
1. Baker, S. C. et al. The External RNA Controls Consortium: a progress report. Nat. Methods 2, 731–4 (2005)