Figure 3.

Comparing bias of leading library prep methods with randomized splint ligation. (A) Normalized read counts of libraries prepared from an equimolar mix of 962 synthetic miRNA are plotted in logarithmic scale. Each miRNA was expected to have a normalized read value of 1 (central dashed line). The upper and lower dashed lines correspond to the interval of 2-fold over- or underrepresented. The total number of miRNA detected with each method is listed below the X-axis label. (B) Percentage of miRNAs falling within 2-fold of their expected values shown as the average of two technical replicates per method. Error bars represent the standard deviation. Letter codes indicate groups that are significantly different from each other with Tukey corrected P < 0.001. (C) Quantification of human brain miRNAs using qPCR compared to each sequencing method. qPCR values are represented as the average Cq of 4 technical replicates while sequencing values are represented as the average number of reads per million for 3–4 technical replicates per method, both are plotted on a logarithmic scale. Linear models were used to plot the line of best fit for each correlation and the R² value of the correlation is printed in the lower left-hand corner of each plot.