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Space Ranger


Loupe

10x Genomics
Visium Spatial Gene Expression

Space Ranger2.0, printed on 11/17/2024

Gene Expression Metrics

Space Ranger pipelines output key metrics in text format. Below are the definitions of the reported metrics.

Metrics definitions for spaceranger count (FF)

The spaceranger count pipeline outputs metrics_summary.csv which contains a number of key metrics about the barcoding and sequencing process.

MetricDescription
Number of Spots Under TissueThe number of barcodes associated with a spot under tissue.
Number of ReadsTotal number of read pairs that were assigned to this library in demultiplexing.
Mean Reads per SpotThe number of reads, both under and outside of tissue, divided by the number of barcodes associated with a spot under tissue.
Mean Reads Under Tissue per SpotThe number of reads under tissue divided by the number of barcodes associated with a spot under tissue.
Fraction of Spots Under TissueThe fraction of the spots under the tissue.
Median Genes per SpotThe median number of genes detected per spot under tissue-associated barcode. Detection is defined as the presence of at least 1 UMI count.
Median UMI Counts per SpotThe median number of UMI counts per tissue covered spot.
Valid BarcodesFraction of reads with barcodes that match the whitelist after barcode correction.
Valid UMIsFraction of reads with valid UMIs; i.e. UMI sequences that do not contain Ns and that are not homopolymers.
Sequencing SaturationThe fraction of reads originating from an already-observed UMI. This is a function of library complexity and sequencing depth. More specifically, this is the fraction of confidently mapped, valid spot-barcode, valid UMI reads that had a non-unique (spot-barcode, UMI, gene).
Q30 Bases in BarcodeFraction of spot barcode bases with Q-score >= 30, excluding very low quality/no-call (Q <= 2) bases from the denominator.
Q30 Bases in RNA ReadFraction of RNA read bases with Q-score >= 30, excluding very low quality/no-call (Q <= 2) bases from the denominator. This is Read 2 for the Visium v1 chemistry.
Q30 Bases in UMIFraction of UMI bases with Q-score >= 30, excluding very low quality/no-call (Q <= 2) bases from the denominator.
Reads Mapped to GenomeFraction of reads that mapped to the genome.
Reads Mapped Confidently to GenomeFraction of reads that mapped uniquely to the genome. If a gene mapped to exonic loci from a single gene and also to non-exonic loci, it is considered uniquely mapped to one of the exonic loci.
Reads Mapped Confidently to Intergenic RegionsFraction of reads that mapped uniquely to an intergenic region of the genome.
Reads Mapped Confidently to Intronic RegionsFraction of reads that mapped uniquely to an intronic region of the genome.
Reads Mapped Confidently to Exonic RegionsFraction of reads that mapped uniquely to an exonic region of the genome.
Reads Mapped Confidently to TranscriptomeFraction of reads that mapped to a unique gene in the transcriptome. The read must be consistent with annotated splice junctions. These reads are considered for UMI counting.
Reads Mapped Antisense to GeneFraction of reads confidently mapped to the transcriptome, but on the opposite strand of their annotated gene. A read is counted as antisense if it has any alignments that are consistent with an exon of a transcript but antisense to it, and has no sense alignments.
Fraction Reads in Spots Under TissueThe fraction of valid-barcode, confidently-mapped-to-transcriptome reads with spot-associated barcodes.
Total Genes DetectedThe number of genes with at least one UMI count in any tissue covered spot.

Metrics definitions for spaceranger count (FFPE)

The spaceranger count for FFPE pipeline outputs metrics_summary.csv which differs from spaceranger count with regard to a few key metrics.

MetricDescription
Reads Mapped to the Probe SetFraction of reads that map with at least one read half to the probe reference.
Reads Mapped Confidently to the Probe SetFraction of reads that map uniquely with both read halves to the probe reference.
Reads Mapped Confidently to the
Filtered Probe Set		Fraction of reads that map uniquely with both read halves to the filtered probe reference. The probe reference is filtered to remove genes/features where one or more of the probes targeting this feature might hybridize and ligate at a non targeted loci. This metric will be None when probe filtering is disabled.
Genes DetectedThe number of unique genes from the filtered probe set with at least one UMI count in any tissue covered spot.
Number of GenesThe number of genes as defined by the probe set.
Number of Genes >= 10 UMIsNumber of genes with at least 10 filtered UMIs from tissue-associated barcodes. These genes are used to calculate per-gene enrichments.

Targeted Spatial Gene Expression metrics

A Targeted Spatial Gene Expression library has additional metrics specific to targeting.

MetricDescription
Analyzed ReadsFraction of the reads used for mapping and analysis. This fraction will be less than 100% when the sequencing depth exceeds 15,000 Mean Reads Under Tissue per Spot. See Targeted Subsampling.
Mean Targeted Reads per SpotThe number of targeted reads, both under and outside of tissue, divided by the number of barcodes associated with a spot under tissue.
Number of Targeted GenesNumber of targeted genes as specified in the input target panel file.
Median Targeted Genes per SpotThe median number of targeted genes detected per tissue-associated barcode. Detection is defined as the presence of at least 1 UMI count. This metric will appear instead of the metric Median Genes per Spot above.
Median Targeted UMI Counts per SpotThe median number of targeted UMI counts per tissue-associated barcode. This metric will appear instead of the metric Median UMI Counts per Spot above.
Targeted Sequencing SaturationThe fraction of targeted reads originating from an already-observed targeted UMI. This is a function of library complexity and sequencing depth. More specifically, this is the fraction of confidently mapped, valid spot-barcode, valid targeted UMI reads that had a non-unique (spot-barcode, UMI, gene). This metric will appear instead of the metric Sequencing Saturation above.
Total Targeted Genes DetectedThe number of targeted genes with at least one UMI count in any spot. This metric will appear instead of the metric Total Genes Detected above.
Number of Targeted GenesNumber of targeted genes as specified in the input target panel file.
Number of Non-Targeted GenesNumber of genes in the reference genome that are not targeted.
Number of Targeted Genes >= 10 UMIsNumber of targeted genes with at least 10 UMIs in spot-associated barcodes. These genes are then considered in the calculation of per-gene enrichments.
Number of Non-Targeted Genes >= 10 UMIsNumber of non-targeted genes with at least 10 UMIs in spot-associated barcodes. These genes are then considered in the calculation of per-gene enrichments.
Number of Enriched Targeted GenesNumber of targeted genes that are classified as enriched as a result of having high mean reads per UMI. Only genes with at least 10 spot-associated UMIs can be enriched. See Targeted Gene Expression Algorithms.
Number of Enriched Non-Targeted GenesNumber of non-targeted genes that are classified as enriched as a result of having high mean reads per UMI. Only genes with at least 10 spot-associated UMIs can be enriched. See Targeted Gene Expression Algorithms..
Mean Reads per UMI per Targeted GeneMean number of reads per UMI per targeted gene, averaged across all targeted genes with at least 10 spot-associated UMIs.
Mean Reads per UMI per Non-Targeted GeneMean number of reads per UMI per non-targeted gene, averaged across all targeted genes with at least 10 spot-associated UMIs.
Reads Mapped Confidently to the
Targeted Transcriptome		Fraction of Reads Mapped Confidently to the Targeted Transcriptome.
Reads Mapped Confidently to the
Non-targeted Transcriptome		Fraction of reads that mapped to a unique gene in the transcriptome. The read must be consistent with annotated splice junctions. These reads are considered for UMI counting. This fraction is computed separately for targeted and non-targeted gene categories.
Fraction of Targeted Genes EnrichedFraction of Targeted Genes with >= 10 spot-associated UMIs that are enriched.
High Fraction of Non-Targeted Genes EnrichedHigh Fraction of Non-Targeted Genes Enriched.