📄 Methods & data

How the results on this site were produced, and where the underlying samples came from.

RNA-seq pipeline

Tumor RNA-seq was processed with the nf-core/oncoanalyser pipeline (Hartwig tooling) in an RNA-only, tumor-only configuration — there is no matched normal and no DNA sequencing in this analysis, so all DNA/germline modules are intentionally disabled.

FASTQ → fastp (trim) → STAR (align) → Isofox (expression + fusions) → CUPPA (tissue of origin)

Isofox produces per-gene and per-transcript expression, gene fusions, and splicing metrics.
CUPPA predicts molecular tissue of origin from the RNA features.
Quality control uses FastQC and alignment metrics aggregated with MultiQC.

GTEx comparison: the same two RNA samples were also reprocessed through the GTEx V11 pipeline (STAR → RSEM → RNA-SeQC) so their expression can be compared directly against GTEx normal tissues and PCAWG tumors — see Expression vs GTEx.

Sequencing quality control

Per-sample RNA-seq quality metrics from Isofox; both tumor samples pass QC.

Sample	QC	Total fragments	Duplicates	Spliced genes	Read length	Median fragment
Left hand (primary)	PASS	41,404,403	23.5%	21,354	76 bp	299 bp
Lung metastasis	PASS	124,808,404	50.5%	26,729	101 bp	133 bp

On the numbers: the lung-metastasis sample has a higher duplicate rate and shorter fragments, consistent with an FFPE / low-input library. Both samples still pass and yield tens of thousands of expressed genes.

Open full MultiQC report ↗

Samples shown on this site

2 of 6 catalogued specimens have RNA-seq reprocessed locally and shown here:

Left hand — primary (SUH-21-014093, FFPE). Fusion: EWSR1::KLF15.
Lung, left lower lobe — metastasis (SP-22-081918-C, Originally Frozen). Fusion: EWSR1::KLF15.

Full sample catalogue

#	Accession	Site	Phase	Type	Diagnosis	Fusion	NGS	Vendor
1	SUH-21-014093	Left hand	Primary / earliest known specimen	FFPE	MEC	EWSR1::KLF15	WES, RNA-seq, single cell	Tempus; NeoGenomics; BostonGene
2	LPS-22-04880	Lung, right lower lobe	Pulmonary metastatic / recurrent lesion	FFPE	MEC	Not yet documented	N/A	N/A
3	CF-01500-2	Lung, left lower lobe	Pulmonary metastatic / recurrent lesion	Cryopreserved	MEC	EWSR1::KLF15	WES, RNA-seq	BGI Genomics
4	SP-22-081918-C	Lung, left lower lobe	Pulmonary metastatic / recurrent lesion	Originally Frozen	MEC	EWSR1::KLF15	RNA-seq	Covance by Labcorp
5	SP-22-081918-A	Lung, right lower lobe	Pulmonary metastatic / recurrent lesion	FFPE	MEC	Not yet documented	WES, RNA-seq, single cell	Tempus; BostonGene
6	SP-22-081918-B	Lung, right upper lobe	Pulmonary metastatic / recurrent lesion	FFPE	MEC	Not yet documented	N/A	N/A

Diagnosis abbreviations are as recorded in the clinical sample sheet. Several specimens have whole-exome sequencing (WES) held at external vendors; those data are not part of this analysis.

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