Flow Documentation

Partek Flow Documentation

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A single-end read that is considered “compatible” would be any read that overlaps the exon 100% as described above in the paragraph on “exonic” reads. For compatible reads, PGS gives raw read counts and RPKM (scaled) counts, while incompatible reads are presented by RPKM counts only (“incompatible RPKM”) and are given in the transcripts spreadsheet (Figure 3). When calculating the RPKM values for incompatible reads, PGS still uses the exon length, as previously described.

If a single-end read includes an exon to exon junction, the region skipped in the read must be consistent with an intron in that transcript. If it is not, the read will be incompatible.

For example, with an alignment that begins at base 100 and has a cigar score of 5M 10N; 5M, the M means alignment Match and the N means skipped bases (junction). To be compatible, bases 100-104 must be exonic, bases 105-115 must be intronic and bases 116-120 must be exonic.

Compatible reads with junction are reported in a separate block of columns in the transcripts spreadsheet (“junction RPKM”).

 

Numbered figure captions
SubtitleText Number of reads per transcript (rows) and sample (columns). “Transcripts” spreadsheet provides raw number of compatible reads as well as scaled number of reads (RPKM) for each sample. In addition, Partek® Genomics Suite™ gives the scaled number of reads for compatible junction reads and for incompatible reads. For definitions, please refer to the text
AnchorNamenum-of-reads

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Paired-End Data Scenarios 

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Considering genes with multiple transcripts, a read can be both counted as compatible for some transcripts, as well as counted incompatible for other transcripts of the same gene (Figure 43). Please note that this concept holds for single-end reads as well.

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Furthermore, all the reads that have at least one alignment contribute to the denominator of the RPKM (millions of _mapped reads_ per sample). Similar to that, the denominator of the RPKM contains all the mapped reads, regardless of whether or not a read is compatible with any transcript.total number of aligned reads.

With respect to the gene-level summarization, user can decide whether or not to consider intronic reads (both single- and paired-end) as compatible with the gene (Figure 5). In the latter case, the entire gene is basically treated as one giant exon. In the case that one end of a paired-end read falls within a gene, and the other end does not, the read will not be included in the gene-level summarization.

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