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Hierarchical clustering of the combined data set clearly separated the two cell lines for all template inputs, including single cells and reference RNA samples.
A second specific analyst added the plasmid DNA in a dedicated template addition area and used the same instruments for all template additions.
Hierarchical clustering based on the top 200 differentially expressed genes from the 10 μg reference data again showed clear separation of the two cell lines for all template inputs, with strong correlation seen between the 10 μg reference differentially expressed data and the 1 ng data (Pearson correlation 0.89), and between the 10 μg reference and the single cell data (Pearson correlation 0.78).
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We also present the analyzed results for all templates with Distance = 1 and 3 (Fig. S6).
Based on these conditions, we again investigated the two requirements for all templates to determine the dynamic range of our digital counting system (Figs 4a,b, and S5).
Next, we optimized parameters and applied these analyses for all templates that contained a wide range of copy numbers, from 20 to 40,000.
There were plateaus for all templates in both type of plots, as a function of the number of random bases and coverage, respectively, suggesting that the selected parameters allow for accurate digital quantification for templates with a high range of copy numbers.
For the number of fixed bases, we used four fixed bases (the 16th, 21st, 24th and 28th fixed bases from the left in the barcode (Table S1) were used for all templates) since the determined number of clusters became close to a constant value at four fixed bases (Figs 3c and S3c).
Independent keys are used for all templates and the key is a one-use key. Figure 2 Difference between the previously proposed image matching and the image matching proposed herein.
For all templates, more RNA was produced using E. coli RNAP compared to T7 RNAP under similar conditions (see Materials and Methods).
Second, for T0447, which is also a three-domain target, the plot revealed that the weighted sum and the whole-chain GDT-TS are about the same, clustering near 90% GDT-TS for all template-based servers.
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