Read summary

Read quality and alignment statistics generated by fastcat bamstats for each input sample. Shown are distributions of read length and quality score, and a summary of alignment outcomes (mapped, unmapped, primary, supplementary) against the reference genome.

ref sample_name total primary secondary supplementary unmapped qcfail duplicate duplex duplex_forming status
SIRV1 control_rep1 12 12 0 0 0 0 0 0 0 Mapped
SIRV2 control_rep1 5 5 0 0 0 0 0 0 0 Mapped
SIRV3 control_rep1 14 14 0 0 0 0 0 0 0 Mapped
SIRV4 control_rep1 11 11 0 0 0 0 0 0 0 Mapped
SIRV5 control_rep1 45 44 0 1 0 0 0 0 0 Mapped
SIRV6 control_rep1 107 104 0 3 0 0 0 0 0 Mapped
SIRV7 control_rep1 8 7 0 1 0 0 0 0 0 Mapped
* control_rep1 3 0 0 0 3 0 0 0 0 Unmapped
ref sample_name total primary secondary supplementary unmapped qcfail duplicate duplex duplex_forming status
SIRV1 control_rep2 12 12 0 0 0 0 0 0 0 Mapped
SIRV2 control_rep2 5 5 0 0 0 0 0 0 0 Mapped
SIRV3 control_rep2 14 14 0 0 0 0 0 0 0 Mapped
SIRV4 control_rep2 11 11 0 0 0 0 0 0 0 Mapped
SIRV5 control_rep2 45 44 0 1 0 0 0 0 0 Mapped
SIRV6 control_rep2 107 104 0 3 0 0 0 0 0 Mapped
SIRV7 control_rep2 8 7 0 1 0 0 0 0 0 Mapped
* control_rep2 3 0 0 0 3 0 0 0 0 Unmapped
ref sample_name total primary secondary supplementary unmapped qcfail duplicate duplex duplex_forming status
SIRV1 treated_rep1 16 16 0 0 0 0 0 0 0 Mapped
SIRV2 treated_rep1 7 7 0 0 0 0 0 0 0 Mapped
SIRV3 treated_rep1 12 12 0 0 0 0 0 0 0 Mapped
SIRV4 treated_rep1 14 14 0 0 0 0 0 0 0 Mapped
SIRV5 treated_rep1 33 33 0 0 0 0 0 0 0 Mapped
SIRV6 treated_rep1 112 110 0 2 0 0 0 0 0 Mapped
SIRV7 treated_rep1 8 8 0 0 0 0 0 0 0 Mapped
* treated_rep1 0 0 0 0 0 0 0 0 0 Unmapped
ref sample_name total primary secondary supplementary unmapped qcfail duplicate duplex duplex_forming status
SIRV1 treated_rep2 16 16 0 0 0 0 0 0 0 Mapped
SIRV2 treated_rep2 7 7 0 0 0 0 0 0 0 Mapped
SIRV3 treated_rep2 12 12 0 0 0 0 0 0 0 Mapped
SIRV4 treated_rep2 14 14 0 0 0 0 0 0 0 Mapped
SIRV5 treated_rep2 33 33 0 0 0 0 0 0 0 Mapped
SIRV6 treated_rep2 112 110 0 2 0 0 0 0 0 Mapped
SIRV7 treated_rep2 8 8 0 0 0 0 0 0 0 Mapped
* treated_rep2 0 0 0 0 0 0 0 0 0 Unmapped

Sample metadata

Metadata for each input sample as parsed from the sample sheet and workflow parameters. Includes the sample alias, barcode, type, and any experimental design columns such as condition or batch that were supplied for differential analysis.

Field Value
barcode barcode01
type test_sample
run_ids ['8c239806e6f576cd17d6b7d532976b1fe830f9c6']
basecall_models []
alias control_rep1
condition control
batch b1
n_primary 197
n_unmapped 3
src_xam None
src_xai None
has_stats True
Field Value
barcode barcode02
type test_sample
run_ids ['8c239806e6f576cd17d6b7d532976b1fe830f9c6']
basecall_models []
alias control_rep2
condition control
batch b2
n_primary 197
n_unmapped 3
src_xam None
src_xai None
has_stats True
Field Value
barcode barcode03
type test_sample
run_ids ['8c239806e6f576cd17d6b7d532976b1fe830f9c6']
basecall_models []
alias treated_rep1
condition treated
batch b1
n_primary 200
n_unmapped 0
src_xam None
src_xai None
has_stats True
Field Value
barcode barcode04
type test_sample
run_ids ['8c239806e6f576cd17d6b7d532976b1fe830f9c6']
basecall_models []
alias treated_rep2
condition treated
batch b2
n_primary 200
n_unmapped 0
src_xam None
src_xai None
has_stats True

Reference and Annotation Checks

Results of compatibility checks between the supplied reference genome and annotation performed before bambu transcript modelling. Build and provider hints are inferred from sequence names and file content to help identify mismatched genome/annotation combinations.

Check Value
Overlapping seqnames 7
Seqnames only in annotation 0
Seqnames only in reference 0
Check Value
Transcript annotation records retained 354
Unstranded records excluded 0
Transcript annotation attributes sanitised 0

Build and Provider Hints

Evidence Hints
Reference build None detected
Annotation build None detected
Reference provider None detected
Annotation provider None detected

Bambu Quality Control

Quality metrics from the bambu transcript discovery and quantification run. Library size statistics show cohort-level summary values to flag large inter-sample variation that could affect CPM normalisation, with the individual per-sample counts listed below. The transcript discovery table shows the transcriptome mode used, the novel discovery rate (NDR) threshold applied, and how many transcripts were present before and after low-count filtering.

Library Size Statistics

Metric Value
Samples analyzed 4
Median library size 198 reads
Min library size 196 reads
Max library size 199 reads
Library size ratio (max/min) 1.02x

Transcript Discovery

Metric Value
Transcriptome mode discover
NDR used automatic
Transcripts before filtering 68
Transcripts after filtering 53
Transcripts removed 15
Median transcripts per sample 48
Unique genes (after filter) 7

Per-Sample Library Sizes

Sample Library Size
control_rep1 199
control_rep2 199
treated_rep1 196
treated_rep2 196

Cohort transcriptome

Summary of the joint cohort bambu transcriptome model. Shows the total number of transcripts and genes in the final model, and the top 500 rows of the transcript abundance count table.

Metric Value
Transcripts 53
Genes 7

Top 500 most abundant transcripts

TXNAME GENEID NDR novelGene novelTranscript txClassDescription readCount relReadCount relSubsetCount txid eqClassById control_rep1 control_rep2 treated_rep1 treated_rep2
SIRV101 SIRV1 False False annotation 1 1 1.0 1.0 0.0 0.0
SIRV102 SIRV1 False False annotation 2 2 1.0 1.0 3.0 3.0
SIRV103 SIRV1 False False annotation 3 3 1.0 1.0 2.0 2.0
SIRV105 SIRV1 False False annotation 4 4 2.0 2.0 3.0 3.0
SIRV109 SIRV1 False False annotation 7 7 7.0 7.0 8.0 8.0
SIRV201 SIRV2 False False annotation 8 8 2.66667 2.66667 0.0 0.0
SIRV202 SIRV2 False False annotation 9 9 1.33333 1.33333 4.0 4.0
SIRV203 SIRV2 False False annotation 10 10 0.0 0.0 2.0 2.0
SIRV204 SIRV2 False False annotation 11 11 0.5 0.5 0.5 0.5
SIRV206 SIRV2 False False annotation 13 13 0.5 0.5 0.5 0.5
SIRV302 SIRV3 False False annotation 15 15 1.54708 1.54708 2.0 2.0
SIRV303 SIRV3 False False annotation 16 14;16 1.70776 1.70776 0.0 0.0
SIRV304 SIRV3 False False annotation 17 17 0.0 0.0 2.5 2.5
SIRV305 SIRV3 False False annotation 18 18 2.49677 2.49677 2.5 2.5
SIRV307 SIRV3 False False annotation 20 20 1.24839 1.24839 0.0 0.0
SIRV308 SIRV3 False False annotation 21 21 2.0 2.0 2.0 2.0
SIRV309 SIRV3 False False annotation 22 22 2.0 2.0 2.0 2.0
SIRV310 SIRV3 False False annotation 23 23 3.0 3.0 1.0 1.0
SIRV403 SIRV4 False False annotation 25 25 4.0 4.0 2.0 2.0
SIRV404 SIRV4 False False annotation 26 26 1.0 1.0 1.0 1.0
SIRV406 SIRV4 False False annotation 28 28 4.0 4.0 4.0 4.0
SIRV408 SIRV4 False False annotation 29 29 1.0 1.0 2.0 2.0
SIRV410 SIRV4 False False annotation 31 31 1.0 1.0 5.0 5.0
SIRV502 SIRV5 False False annotation 33 33 2.31273 2.31273 1.34771 1.34771
SIRV504 SIRV5 False False annotation 35 35 1.0 1.0 1.0 1.0
SIRV505 SIRV5 False False annotation 36 36 5.27469 5.27469 4.88546 4.88546
SIRV506 SIRV5 False False annotation 37 32;33;36;37;39;41 7.37957 7.37957 6.43348 6.43348
SIRV507 SIRV5 False False annotation 38 38 11.0 11.0 7.0 7.0
SIRV508 SIRV5 False False annotation 39 39 3.95601 3.95601 1.62849 1.62849
SIRV509 SIRV5 False False annotation 40 40 2.58188 2.58188 0.0 0.0
SIRV510 SIRV5 False False annotation 41 41 2.21383 2.21383 2.45185 2.45185
SIRV511 SIRV5 False False annotation 42 42 3.28129 3.28129 2.25301 2.25301
SIRV512 SIRV5 False False annotation 43 43 4.0 4.0 3.0 3.0
SIRV601 SIRV6 False False annotation 44 44 1.14286 1.14286 1.14856 1.14856
SIRV602 SIRV6 False False annotation 45 45 24.0 24.0 21.0 21.0
SIRV603 SIRV6 False False annotation 46 46;53 1.0 1.0 1.08914 1.08914
SIRV604 SIRV6 False False annotation 47 47 6.85714 6.85714 5.85144 5.85144
SIRV605 SIRV6 False False annotation 48 48;55 10.0 10.0 10.0 10.0
SIRV606 SIRV6 False False annotation 49 44;47;48;49;53;55 4.88316 4.88316 6.53036 6.53036
SIRV607 SIRV6 False False annotation 50 50 7.0 7.0 4.0 4.0
SIRV608 SIRV6 False False annotation 51 44;45;51;55 4.0 4.0 3.0 3.0
SIRV609 SIRV6 False False annotation 52 47;48;52;55 18.0 18.0 22.0 22.0
SIRV610 SIRV6 False False annotation 53 53 3.0 3.0 3.0 3.0
SIRV611 SIRV6 False False annotation 54 54 2.0 2.0 2.0 2.0
SIRV614 SIRV6 False False annotation 57 57 10.0 10.0 10.0 10.0
SIRV615 SIRV6 False False annotation 58 44;45;51;55;56;58 10.0 10.0 13.91086 13.91086
SIRV616 SIRV6 False False annotation 59 59 4.11684 4.11684 3.46964 3.46964
SIRV617 SIRV6 False False annotation 60 60 0.0 0.0 2.0 2.0
SIRV702 SIRV7 False False annotation 63 63 3.0 3.0 3.0 3.0
SIRV703 SIRV7 False False annotation 64 64 3.0 3.0 3.0 3.0
SIRV704 SIRV7 False False annotation 65 65 2.0 2.0 1.0 1.0
SIRV706 SIRV7 False False annotation 67 67 0.0 0.0 0.5 0.5
SIRV708 SIRV7 False False annotation 68 68 0.0 0.0 0.5 0.5

Per-sample transcriptomes

Per-sample transcript and gene count summaries derived from the individual bambu quantification runs. Each tab shows the number of transcripts and genes detected in that sample after filtering. These can be used to spot samples with unusually low transcript detection compared to the rest of the cohort.

Metric Value
Transcripts 48
Genes 7
Metric Value
Transcripts 48
Genes 7
Metric Value
Transcripts 48
Genes 7
Metric Value
Transcripts 48
Genes 7

SQANTI3 classification

Structural classification of transcript isoforms by SQANTI3. Each transcript is assigned a category based on how its splice junctions and exon structure compared to the reference annotation. The table shows the count of transcripts in each category per sample and for the whole cohort.

Sample Full splice match Incomplete splice match Novel in catalog Novel not in catalog Antisense Genic intron Genic Intergenic
control_rep1 48 0 0 0 0 0 0 0
control_rep2 48 0 0 0 0 0 0 0
treated_rep1 48 0 0 0 0 0 0 0
treated_rep2 48 0 0 0 0 0 0 0
cohort 53 0 0 0 0 0 0 0

Full splice match: Reference and query isoforms have the same number of exons and all internal junctions agree.
 Incomplete splice match: Query isoform has fewer 5′ exons than the reference, with matching internal junctions.
 Novel in catalog: No full or incomplete splice match, but uses a combination of known donor/acceptor splice sites.
 Novel not in catalog: No full or incomplete splice match, with at least one unannotated donor or acceptor splice site.
 Antisense: No same-strand reference overlap, but antisense to an annotated gene.
 Genic intron: Query isoform is fully contained within an annotated intron.
 Genic: Query isoform overlaps introns and exons.
 Intergenic: Query isoform lies in an intergenic region.

Differential Analysis Quality Control

Quality control summary for the differential expression analyses. Shows the experimental design and the number of samples per group. For each contrast, the statistical methods chosen by DESeq2 and DEXSeq are reported , including the dispersion estimation strategy (parametric or gene-wise fallback) and size factor normalisation method, alongside the analysis status and count of significant hits. Any warnings about low sample numbers, dispersion fallbacks, dropped covariates, or analysis failures are highlighted here.

Experimental Design

Parameter Value
Total samples 4
Condition column condition
Reference level control
Covariates batch
Number of contrasts 1

Sample Sizes per Group

Group Samples Status Note
control 2 ⚠️ Low power
treated 2 ⚠️ Low power

Statistical Methods & Warnings

Contrast DESeq2 size factors DESeq2 dispersion DEXSeq size factors DEXSeq dispersion DEXSeq covariates dropped DGE status DTU status
condition_treated_vs_control ratio gene-wise (fallback) ratio parametric none SUCCESS FAILED

Results Summary by Contrast

Contrast Samples DGE Status DGE Significant (FDR<0.05) DGE Up DGE Down DTU Status DTU Genes (q<0.05)
condition_treated_vs_control 2 vs 2 SUCCESS 0 0 0 FAILED N/A

Quality Warnings Summary

Warning Type Details
Sample Size Some groups have n<3 (recommended minimum)
Gene-wise Dispersion Fallback DESeq2 (1 contrasts)
DTU Failure 1 contrasts failed

Differential gene expression

Differential gene expression results from DESeq2. The heatmap, PCA plot, and sample distance matrix are derived from CPM-normalised counts and give an overview of sample clustering relative to the experimental conditions. Each contrast tab shows a results table of genes ranked by adjusted p-value and a volcano plot highlighting significantly up- and down-regulated genes.


(Left) Hierarchical clustering heatmap of the top 150 most variable gene across all samples. Rows represent gene (Z-scored and log2 transformed fold changes), columns represent samples. Dendrograms show clustering of both genes and samples. (Middle) Principal component analysis (PCA showing the first two principal components) of sample gene expression profiles. Each point represents a sample, coloured by condition. Samples that cluster together have similar overall expression profiles. (Right) Sample-to-sample Euclidean distance matrix calculated from log2-transformed fold change values. Lower values (darker blue) indicate more similar expression profiles between samples.

Note: DTU analysis may be underpowered (n=4, recommend n>=6 with >=3 per group)

GENEID newGeneClass baseMean log2FoldChange lfcSE stat pvalue padj
SIRV5 annotation 39.118 -0.501 0.234 -2.142 0.032 0.225
SIRV1 annotation 13.965 0.362 0.390 0.928 0.353 0.689
SIRV2 annotation 5.983 0.432 0.597 0.723 0.470 0.689
SIRV3 annotation 13.021 -0.276 0.401 -0.687 0.492 0.689
SIRV4 annotation 12.474 0.294 0.411 0.716 0.474 0.689
SIRV6 annotation 107.981 6.632e-08 0.139 4.776e-07 1.000 1.000
SIRV7 annotation 8.001 -0.053 0.510 -0.105 0.917 1.000
Table showing the top 500 genes sorted by adjusted p-value.


Gene expression volcano Plot

Differential transcript usage

Differential transcript usage results from DEXSeq. Unlike DGE, DTU tests whether individual transcripts change their proportional contribution to total gene expression between conditions , i.e. isoform switching , rather than testing for changes in total gene abundance. The heatmap, PCA, and sample distance matrix use CPM-normalised counts. Each contrast tab shows a results table ranked by adjusted p-value and a volcano plot.


(Left) Hierarchical clustering heatmap of the top 150 most variable transcript across all samples. Rows represent transcript (Z-scored and log2 transformed fold changes), columns represent samples. Dendrograms show clustering of both genes and samples. (Middle) Principal component analysis (PCA showing the first two principal components) of sample transcript expression profiles. Each point represents a sample, coloured by condition. Samples that cluster together have similar overall expression profiles. (Right) Sample-to-sample Euclidean distance matrix calculated from log2-transformed fold change values. Lower values (darker blue) indicate more similar expression profiles between samples.

Empty results indicate analysis failure, not 'no DTU detected'.

No DTU results available for this contrast.

Software versions

Name Version
minimap2 2.28-r1209
samtools 1.23.1
gffread 0.12.9
bambu 3.12.1
DESeq2 1.50.2
DEXSeq 1.56.0
SQANTI3 5.5.1
modkit 0.6.3

Workflow parameters

Key Value
fastq test_data/smoke/de
bam None
sample None
sample_sheet test_data/smoke/sample_sheet_de.csv
out_dir wf-transcriptomes
igv False
ref_genome test_data/smoke/reference.fa
ref_annotation test_data/smoke/annotation.gtf
transcriptome_mode discover
direct_rna False
de_analysis True
condition_column condition
covariates batch
reference_level control
analyse_unclassified False
analyse_fail False
mod_codes None
force_alignment False
ndr None
sqanti_skip_orf True
store_dir wf-transcriptomes/store_dir