Subsections

QIAseq miRNA Quantification outputs

The tool will output expression tables. The "Grouped on mature" table has a row for each mature miRNA. The same mature miRNA may be produced from different precursor miRNAs. The "Grouped on seed" table has a row for each seed sequence. The same seed sequence may be found in different mature miRNAs. If a custom database was provided, a "Grouped on custom database" will be added to the output folder.

Grouped on seed

(Image rna_seeds_table_16_n_p)

In this expression table, there is a row for each seed sequence (figure 14.26).

Image mirnaseed
Figure 14.26: Expression table grouped on seed.

This table contains the following information:

Grouped on mature

(Image annotate_small_rna)

In this table, there is a row for each mature miRNA in the database, including those for which the expression is zero (figure 14.27). Double click on a row to open a unique reads alignment (seen at the bottom of figure 14.27). Unique reads result from collapsing identical reads into one. The number of reads that are collapsed into a unique read is indicated in parentheses to the right of the miR name of the unique mature read. The alignment shows all possible unique reads that have aligned to a specific miRNA from the database. Mismatches to the mature reference are highlighted in the alignment and recapitulated in their name as explained in Naming isomiRs.

Image mirnamature
Figure 14.27: Expression table grouped on mature, with a view of a unique reads alignment.

This table contains the following information:

Grouped on custom database

(Image annotate_small_rna)

In this table, there is a row for each mature smallRNA in the database, including those for which the expression is zero (figure 14.28). Double click on a row to open a unique reads alignment (seen at the bottom of figure 14.28). Unique reads result from collapsing identical reads into one. The number of reads that are collapsed into a unique read is indicated in parentheses to the right of the miR name of the unique mature read. The alignment shows all possible unique reads that have aligned to a specific miRNA from the database. As with the table Grouped on mature, mismatches to the reference are highlighted in the alignment and recapitulated in their name as explained in Naming isomiRs.

Image mirnacustom
Figure 14.28: Expression table grouped on custom database, with a view of a unique reads alignment.

This table contains the following information:

Reports and discarded reads

The workflow also outputs reports:

The quantification report contains the following main sections:

It is later possible to combine all reports issued for one sample using the Create Combined miRNA Report tool (see http://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Create_Combined_miRNA_Report.html.


Naming isomiRs

The names of aligned sequences in mature groups adhere to a naming convention that generates unique names for all isomiRs. This convention is inspired by the discussion available here: http://github.com/miRTop/incubator/blob/master/isomirs/isomir_naming.md

Deletions are in lowercase and there is a suffix s for 5' deletions (figure 14.29):

Image mirnanamingdel
Figure 14.29: Naming of deletions.

Insertions are in uppercase and there is a suffix s for 5' insertions (figure 14.30):

Image mirnanamingins
Figure 14.30: Naming of insertions.

Note that indels within miRNAs are not supported.

Mutations (SNVs) are indicated with reference symbol, position and new symbol. Consecutive mutations will not be merged into MNVs. The position is relative to the reference, so preceding (5') indels will not offset it (figure 14.31):

Image mirnanamingmut
Figure 14.31: Naming of mutations.

Deletions followed by insertions will be annotated as shown in figure 14.32:

Image mirnanamingdelinse
Figure 14.32: Naming of deletions followed by insertions.

If a sequence maps to multiple miRBase entries or to multiple entries in a custom database, we will add the suffix `ambiguous' to its name. This can happen when multiple species are selected, as they will often share the same miRBase (or other reference) sequence, or when a read does not map perfectly to any miRBase entry, but is close to two or more entries, distinguished by just one SNV, for example.