Cogent NGS Analysis Pipeline notices

The issues described below for the Cogent NGS Analysis Profiler are ones that have been seen or reported. If you encounter other errors using CogentAP or are unable to resolve the problem using the suggestions below, please:

Capture a screenshot of the text of the error you see on the screen.
Gather the relevant log file(s). Refer to the FAQ entry below for more detailed information about the logs.
Send the screenshot and gathered log file(s) to Technical Support, along with a brief description of the issue.

Software issues FAQs

List & description of log files

Problem area	Log filename	Directory location
Installation	CogentAP_install.log	`%COGENT_AP_HOME`
Demuxer	demux_demuxer.log	[Configured demux output folder] (see Figures 1 and 2)
Analyzer	analysis_analyzer.log	[Configured analysis output folder] (see Figures 1 and 3)

Data analysis FAQs

Columns in Stats file

The tables below document all potential columns that might display in a Cogent NGS Analysis Pipeline Stats file; they also represent the column names accepted for input into Cogent NGS Discovery Software.

Columns that will be present in all Stats files output by CogentAP (input workflow agnostic).

Column name	Description
Barcode	Detected barcodes. This value will usually be the sample name from the well-list or well-list-like file, but there are three exceptions, documented in the table below.
Sample	Sample names described in sample description file.
Barcoded_Reads	Number of reads after demultiplexing.
Trimmed_Reads	Number of remained reads after trimming.
Unmapped_Reads	Number of reads not mapped to genome.
Mapped_Reads	Number of reads mapped to genome.
Multimapped_Reads	Number of reads mapped to multiple genomic locations.
Uniquely_Mapped_Reads	Number of reads mapped to one genomic location. These reads are used for counting.
Exon_Reads	Number of reads assigned to an exonic region.
Ambiguous_Exon_Reads	Number of reads assigned to exonic regions of multiple genes.
Intron_Reads	Number of reads assigned to an intronic region.
Ambiguous_Intron_Reads	Number of reads assigned to intronic regions of multiple genes.
Gene_Reads	Number of reads assigned to a gene region (exon + intron).
Intergenic_Reads	Number of reads assigned to an intergenic region.
No_of_Genes	Number of detected genes.
Mitochondrial_Reads	Number of reads assigned to mitochondrial chromosome.
Ribosomal_Reads	Number of reads assigned to a ribosomal gene.

The "Barcode" column, in addition to the samples named in the well-list or well-list-like file, will also have three additional rows, which are described in the following table.

Barcode field value	Description
Short	Number of reads containing N in barcode or having shorter length than barcode.
Unselected	Number of reads having a barcode included in Chip’s description, but not included in sample description file.
Undetermined	Number of reads having undetermined barcode.

Additional columns in the Stats file for 3′ DE analysis with UMIs on ICELL8 system.

The table below lists additional columns that will be present in the Stats file when the input FASTQ files result from the ICELL8 3′ DE for UMI Reagent Kit (Cat # 640005) workflow on the ICELL8 Single-Cell System (Cat # 640000).

Column name	Description
No_of_UMIs	Number of UMI variations detected after demultiplexing.
Exon_nUMIs	Number of deduplicated reads assigned to an exonic region. Deduplication is done by UMI.
Intron_nUMIs	Number of deduplicated reads assigned to an intronic region. Deduplication is done by UMI.
Gene_nUMIs	Number of deduplicated reads assigned to a gene region (exon + intron). Deduplication is done by UMI.

Additional columns in Stats file for the SMARTer Stranded Total RNA-Seq Kit v3- Pico Input Mammalian protocol.

The table below lists additional columns that will be present in the Stats file when the input FASTQ files result from the SMARTer Stranded Total RNA-Seq Kit v3- Pico Input Mammalian protocol.

Column name	Description
No_of_UMIs	Number of UMI variations detected after demultiplexing.
Exon_nUMIs	Number of deduplicated reads assigned to an exonic region. Deduplication is done by UMI.
Exon_nUSSs	Number of deduplicated reads assigned to an exonic region. Deduplication is done by Unique Start&Stop Site (USS).
Exon_nUMIs_USSs	Number of deduplicated reads assigned to an exonic region. Deduplication is done by both UMI and USS.
Intron_nUMIs	Number of deduplicated reads assigned to an intronic region. Deduplication is done by UMI.
Intron_nUSSs	Number of deduplicated reads assigned to an intronic region. Deduplication is done by Unique Start&Stop Site (USS).
Intron_nUMIs_USSs	Number of deduplicated reads assigned to an intronic region. Deduplication is done by both UMI and USS.
Gene_nUMIs	Number of deduplicated reads assigned to a gene region (exon + intron). Deduplication is done by UMI.
Gene_nUSSs	Number of deduplicated reads assigned to a gene region (exon + intron). Deduplication is done by Unique Start&Stop Site (USS).
Gene_nUMIs_USSs	Number of deduplicated reads assigned to a gene region (exon + intron). Deduplication is done by both UMI and USS.
Strand_Specificity	Ratio of reads detected as correct strand after mapping to genome.

RNA-seq

Cogent NGS Analysis Pipeline

Analyze RNA-seq data generated from ICELL8 system applications.

Cogent NGS Discovery Software

Visualize RNA-seq data in UMAP or t-SNE charts from the output of the Cogent NGS Analysis Pipeline.

SMART-Seq DE3 Demultiplexer

Demultiplex sequencing data from the SMART-Seq v4 3′ DE Kit into sorted read data files.

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