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If the Require junction reads to match introns check button is selected, only junction reads that overlap with exonic regions and match the skipped bases of an intron in the transcript will be included in the calculation. Otherwise, as long as the reads overlap within the exonic region, they will be counted. Detailed information about read compatibility can be found in the Understanding Reads tutorial.

Some library preparations reverse transcribe the mRNA into double stranded cDNA, thus losing strand information. In this case, the total transcript count will include all the reads that map to a transcript location. Others will preserve the strand information of the original transcript by only synthesizing the first strand cDNA.  Thus, only the reads that have sense compatibility with the transcripts will be included in the calculation. We recommend verifying with the data source how the NGS library was prepared to ensure correct option selection.

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In the annotation file, there might be multiple features in the same location, or one read might have multiple alignments, so the read count of a feature might not be an integer. Our white paper on the Partek E/M algorithm has more details on Partek’s implementation the E/M algorithm initially described by Xing et al. [1]

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In sample histogram, each line represents a sample and the range of read counts are divided into 20 bins. Clicking on a sample in the legend will hide the line for that specific sample. Hovering over each circle displays detailed information about the sample and that specific bin (Figure 9). The information includes:

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