QIAGEN Bioinformatics Manuals

Data QC and Remove Background Reads

The Data QC and Remove Background Reads workflow performs trimming of reads, creates a QC report and cleans the dataset from background DNA, leaving back only the reads that match the reference genome(s).

To run the Data QC and Remove Background Reads workflow, go to:

You can select only one read file to analyze (figure 3.11). Alternatively, multiple files can be run using the batch mode.

Image clean_1
Figure 3.11: Select the reads. Use Batch mode if you have more than one set of reads.

In the "Trim Sequences" dialog, you can specify a trim adapter list and set up parameters if you would like to trim your sequences from adapters (figure 3.12).

Image clean_2_wf
Figure 3.12: You can choose to trim adapter sequences from your sequencing reads.

The parameters that can be set are:

Quality limit: defines the minimal value of the Phred score for which bases will not be trimmed.
Trim adapter list: the adapter sequences to trim (if any).

In the Taxonomic Profiling dialog, select the "Species of interest taxpro index" you will use to map the reads (figure 3.13). Here, you can also choose to "Filter background reads". You must then specify the "Background taxpro index" (in the case of human microbiota, the Homo sapiens GRCh38 for example). The reference database can be obtained by using the Download Curated Microbial Reference Database tool (Download Curated Microbial Reference Database) or Download Custom Microbial Reference Database tool (Download Custom Microbial Reference Database). The host index and (if using the custom downloader) the microbial reference index are built with the Create Taxonomic Profiling Index tool (Create Taxonomic Profiling Index).

Image clean_2
Figure 3.13: Select the reference databaes, and potentially a background taxpro index to remove possible contamination.

The workflow will output three sequence lists:

Trimmed, cleaned reads: trimmed reads mapping to the species of interest taxpro index of reference genome(s).
Background reads: reads mapping to the background taxpro index.
Unmapped reads: reads not mapping to the species of interest or the background taxpro index.

In addition, it generates three reports: a trimming report, a graphical QC report and a supplementary QC report. All of these should be inspected in order to determine whether the quality of the sequencing reads and the trimming are acceptable. For a detailed description of the QC reports and indication on how to interpret the different values, see http://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=QC_Sequencing_Reads.html.

For the trimming report, see http://resources.qiagenbioinformatics.com/manuals/clcgenomicsworkbench/current/index.php?manual=Trim_output.html.

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Data QC and Remove Background Reads