BulkSeq Studio 0.16.0

Revised 0.16.0 build (2 Jul 2026): two additive revisions were folded into the binaries below. The self-contained HTML results report was redesigned (branded single file, zoomable SVG figures, separate up- and down-regulated gene tables, functional-enrichment tables, and per-step runtimes), the figures step now also produces separate up- and down-regulated top-DEG heatmaps, and Check Environment probes every optional pipeline route (Trim Galore, Trimmomatic, SortMeRNA, RiboDetector, FastQ Screen, RSeQC, UCSC utilities) and the R analysis packages. The default STAR → featureCounts → DESeq2 analysis path is byte-identical to the original 0.16.0 (the Fusarium graminearum baseline reproduces exactly at 2,723 up / 2,478 down).

BulkSeq Studio 0.16.0 — a large additive feature release. The default STAR → featureCounts → DESeq2 analysis path is byte-identical to 0.15.x (the Fusarium graminearum baseline reproduces exactly at 2,723 up / 2,478 down), so existing results are unchanged; every new capability below is opt-in.

Added

• Alternative differential-expression engines. Alongside DESeq2 (default), RNA-seq counts can be tested with limma-voom or edgeR (quasi-likelihood F-test). All three emit the same result schema, so enrichment, networks, and figures are identical regardless of engine. Concordance with DESeq2 is high (log2FC Spearman 0.997–1.000, top-DEG Jaccard 0.94–0.97, 100% direction agreement; benchmark B14).
• Alternative read trimmers. Selectable fastp (default), Trim Galore, or Trimmomatic, each with its own parameters in the GUI.
• Alternative rRNA removal. SortMeRNA (default, reference-based) or RiboDetector (reference-free, CPU, no database).
• Contamination screening. Optional FastQ Screen step maps a read subsample against a panel of reference genomes.
• Single-end FASTQ support. Single-end libraries now run the full alignment route (trimming, rRNA filtering, STAR/HISAT2/Salmon, featureCounts). Mixed single/paired layouts are rejected with guidance.
• GSVA pathway activity. Optional per-sample gene-set activity scoring from your own custom gene sets (organism-safe), with a heatmap.
• RSeQC alignment QC. Optional read-distribution and gene-body-coverage reports (genome-BAM routes).
• Self-contained HTML report. A single-file results_report.html inlining figures, top genes, enrichment, and provenance; reachable from the GUI.
• Guided design/covariate builder. A dialog composes the DESeq2 design formula from your metadata columns (batch/covariate adjustment) without typing R.
• Advanced parameters panel. A collapsible per-tool section exposing the important parameters of each tool, with the validated defaults pre-filled.

Changed

• Pipeline environments gained edger, trim-galore, cutadapt, pigz, trimmomatic, fastq-screen, bowtie2, ribodetector, gsva, rseqc, and the UCSC gtfToGenePred/genePredToBed utilities; the pinned lock was regenerated (CPU ONNX runtime, no CUDA).
• Validation extended to a human dataset (airway smooth muscle ± dexamethasone, GRCh38/Salmon): the pipeline recovers the canonical glucocorticoid signature (FKBP5, ZBTB16, KLF15, SPARCL1 up; VCAM1 down) — benchmark B15.

Removed

• htseq-count dropped as a quantifier option (featureCounts, STAR gene counts, and Salmon/tximport cover the same ground).

Downloads

• Windows installer: BulkSeqStudio-Setup-0.16.0.exe — per-user install, no admin rights.
• Windows portable: BulkSeqStudio-Portable-0.16.0.zip — unzip and run.
• Linux AppImage: BulkSeqStudio-0.16.0-x86_64.AppImage — chmod +x and run (built on Ubuntu 24.04; needs glibc 2.39+).
• Linux portable: BulkSeqStudio-linux-0.16.0.tar.gz — extract and run the bundled launcher.

The validation benchmark archive (families B1–B15) is on Zenodo: https://doi.org/10.5281/zenodo.20955660

Per-asset SHA-256 checksums are shown next to each file in the Assets section below.

BulkSeq Studio 0.15.2

0.15.2

Fixed

• gffread / salmon / hisat2 not found in Snakemake shell rules. The make_transcriptome, salmon_index, and hisat2_index rules now explicitly prepend ${MAMBA_ROOT_PREFIX}/envs/bulkseq/bin to PATH at the start of their shell commands. Snakemake's subprocess environment does not reliably inherit the micromamba-activated PATH in all configurations; this fixes the "gffread is not installed" error even when the binary is present in the env.

BulkSeq Studio 0.15.1

0.15.1

Fixed

• Workflow Settings tab scaling. Form fields (dropdowns, spinboxes, line edits) now expand to fill the full available width at any window or monitor size. The Qt default field-growth policy left Preferred-policy widgets like QComboBox at a narrow hint with empty space to the right on wide windows.
• Setup readiness: Salmon/HISAT2 route tools warning. When salmon, gffread, or hisat2 are absent from the WSL bulkseq env, the Setup tab's recommended actions now explicitly call this out and direct you to Install/Repair Core WSL Env. Previously a machine missing gffread (needed by the Salmon route's transcriptome step) would show "Setup is ready" while Salmon runs would fail with a runtime error.

BulkSeq Studio 0.15.0

BulkSeq Studio 0.15.0

Two new analysis options. Both are additive — the default pipeline (featureCounts, built-in GO/KEGG enrichment) is unchanged, and the F. graminearum DESeq2 baseline still reproduces exactly.

Added

• STAR gene-counts quantifier. With the STAR aligner, the Quantifier control is now a real choice: STAR_GeneCounts takes gene counts from STAR's own --quantMode GeneCounts output (no separate counting pass), strand-matched to the run's inferred strandedness, instead of running featureCounts. The counts converge on the same matrix the rest of the pipeline expects — validated at Pearson r ≈ 0.998 (unstranded) to 1.000 (reverse-stranded) against featureCounts on the same BAMs. featureCounts stays the default; HISAT2 uses featureCounts and Salmon uses tximport.
• Custom gene-set enrichment. Supply your own gene sets — a GMT and/or an id→term annotation table, with an optional background gene list for the over-representation universe — to run a clusterProfiler ORA + GSEA alongside the built-in GO/KEGG, producing custom ORA/GSEA tables and a dotplot. It needs no Bioconductor OrgDb, so it works even where the built-in GO route is skipped (e.g. most fungi). The gene IDs must use the run's identifier format; a namespace mismatch is flagged rather than returned as a silent empty result. The built-in GO/KEGG enrichment is unchanged.

Downloads

Windows:

• BulkSeqStudio-Setup-0.15.0.exe — installer
• BulkSeqStudio-Portable-0.15.0.zip — unzip and run BulkSeqStudio.exe, no install

Linux (x86-64, glibc 2.39+):

• BulkSeqStudio-0.15.0-x86_64.AppImage — mark executable and run
• BulkSeqStudio-linux-0.15.0.tar.gz — extract and run BulkSeqStudio

BulkSeq Studio 0.14.2

BulkSeq Studio 0.14.2

Improves the genes-of-interest (focused custom-gene) analysis.

Changed

• Genes of interest: identifier-mismatch flag and clearer guidance. Paste a list of genes and BulkSeq Studio produces a focused z-scored heatmap, per-condition expression plots, a normalized-counts table, and — when PPI seeding is set to the gene list — a STRING network for just those genes. The list is matched to the run's genes by locus tag, Ensembl/RefSeq ID, or gene symbol. When few or none match (usually because the IDs are in a different format than the run uses, e.g. gene symbols pasted into a locus-tag run), the genes-of-interest report now leads with a clear warning and shows examples of the run's actual ID format so the list can be corrected. The Genes of Interest tab states the format requirement.

Downloads

Windows:

• BulkSeqStudio-Setup-0.14.2.exe — installer
• BulkSeqStudio-Portable-0.14.2.zip — unzip and run BulkSeqStudio.exe, no install

Linux (x86-64, glibc 2.39+):

• BulkSeqStudio-0.14.2-x86_64.AppImage — mark executable and run
• BulkSeqStudio-linux-0.14.2.tar.gz — extract and run BulkSeqStudio

BulkSeq Studio 0.14.1

BulkSeq Studio 0.14.1

Maintenance release: removes dead configuration scaffolding so the config no longer advertises behavior the pipeline does not perform. No change to results or to any working configuration.

Changed

• Removed unused config fields that no workflow rule or app code read: workflow.repair_pairs (BBMap repair, never implemented), workflow.differential_expression (the edgeR / limma-voom values were never wired — DESeq2 is selected by input mode), sortmerna.enabled (rRNA filtering is gated on workflow.rrna_filtering), and the STAR overrides outSAMtype / quantMode / sjdb_overhang / genomeSAindexNbases (the indexing rule computes these itself). Existing project configs that still contain these keys load unchanged — the stale keys are ignored.
• Removed the orphaned STAR block in tool_defaults.yaml and a progress-label entry for a "repair" rule that does not exist; updated the README so the feature list shows SortMeRNA rRNA filtering as implemented.

Downloads

Windows:

• BulkSeqStudio-Setup-0.14.1.exe — installer
• BulkSeqStudio-Portable-0.14.1.zip — unzip and run BulkSeqStudio.exe, no install

Linux (x86-64, glibc 2.39+):

• BulkSeqStudio-0.14.1-x86_64.AppImage — mark executable and run
• BulkSeqStudio-linux-0.14.1.tar.gz — extract and run BulkSeqStudio

BulkSeq Studio 0.14.0

BulkSeq Studio 0.14.0

A full audit of the GUI controls and analysis scripts (multi-agent, adversarially verified) found no scientific-validity issues — the DESeq2 baseline reproduces exactly — and turned up two GUI controls that looked active but did nothing. Both now work.

Added

• Skip-trimming toggle. The "fastp trimming" checkbox is now honored: unchecking it skips fastp entirely and sends the raw reads straight to the aligner (and to rRNA filtering, if that is on). Previously the box only changed the runtime estimate while fastp always ran. FastQC-before/after and the MultiQC inputs follow the same gating. Leave it on unless your reads are already trimmed.
• GFF3 annotation support. The annotation Format selector (gtf / gff3) is now honored: a GFF3 annotation is converted to GTF with gffread before indexing and counting, so STAR/HISAT2/Salmon and featureCounts get the GTF they expect. Previously selecting gff3 did nothing and a real GFF3 file was silently parsed as GTF (wrong or empty counts). The GTF path is unchanged.

Validation

The default (trimming-on, GTF) path is byte-identical: the F. graminearum DESeq2 baseline still reproduces 2723 up / 2478 down. Trimming-off was run end to end (raw reads → aligner → counts), the trimming-off + rRNA-on combination routes raw reads into SortMeRNA, and a GFF3→GTF round trip is count-identical to the original GTF (featureCounts assigns the same reads to the same genes).

Downloads

Windows:

• BulkSeqStudio-Setup-0.14.0.exe — installer
• BulkSeqStudio-Portable-0.14.0.zip — unzip and run BulkSeqStudio.exe, no install

Linux (x86-64, glibc 2.39+):

• BulkSeqStudio-0.14.0-x86_64.AppImage — mark executable and run
• BulkSeqStudio-linux-0.14.0.tar.gz — extract and run BulkSeqStudio

BulkSeq Studio 0.13.0

BulkSeq Studio 0.13.0

Added

• rRNA filtering with SortMeRNA. The "rRNA filtering" workflow option is now implemented (it was previously a checkbox that did nothing). When enabled, trimmed reads are filtered against the SortMeRNA rRNA database before alignment, on all three aligner routes (STAR, HISAT2, Salmon): the reference is downloaded and indexed once per project, then each sample is filtered in its own working directory and the non-rRNA reads feed the aligner. The per-sample SortMeRNA log (rRNA %) is added to the MultiQC report, and sortmerna is now part of the core environment. A custom reference can be set via sortmerna.database (a local FASTA, a FASTA URL, or a database tarball URL); the default is smr_v4.3_default_db.

Fixed

• Rule guards added in 0.12.2 could abort their own rules. The command -v … || { … } guards in make_transcriptome, salmon_index, and hisat2_index used unescaped braces, which Snakemake parses as format fields, raising a NameError and stopping the Salmon and HISAT2 routes even when the tool was present. The braces are now escaped, so those routes run. (The STAR route was never affected.)

Upgrading

The rRNA option needs sortmerna, now in the core environment: open Check Environment → Install / repair core environment (on Linux, update from workflow/envs/bulkseq_core.yaml). Enabling rRNA filtering downloads a ~150 MB reference and builds a multi-GB index once per project.

Downloads

Windows:

• BulkSeqStudio-Setup-0.13.0.exe — installer
• BulkSeqStudio-Portable-0.13.0.zip — unzip and run BulkSeqStudio.exe, no install

Linux (x86-64, glibc 2.39+):

• BulkSeqStudio-0.13.0-x86_64.AppImage — mark executable and run
• BulkSeqStudio-linux-0.13.0.tar.gz — extract and run BulkSeqStudio

BulkSeq Studio 0.12.3

BulkSeq Studio 0.12.3

Superseded by 0.13.0. This release inherits the 0.12.2 rule-guard bug (unescaped braces parsed by Snakemake as a format field), which aborts the Salmon and HISAT2 routes with a NameError even when the environment is correct. Use 0.13.0 or newer for those aligners. The STAR route and the fixes below are unaffected.

Patch release. Two dependency-coverage gaps found by auditing the rest of the pipeline for the same class as the 0.12.2 aligner-tool fix.

Fixed

• DESeq2 log-fold-change shrinkage could fail with a missing-package error. run_deseq2.R falls back to lfcShrink(type="ashr") for contrasts apeglm cannot shrink, and ashr is also selectable via deseq2.shrinkage_method, but the ashr R package was in no environment profile. A config that requested ashr (or a default apeglm run that hit the contrast fallback) aborted after the model fit. r-ashr is now in the full environment and the pinned lock, and deseq2.shrinkage_method is restricted to apeglm, ashr, or normal so an unsupported value is rejected when the config loads rather than mid-run. Default (apeglm) runs are unchanged.
• GO enrichment for yeast, Arabidopsis, C. elegans, and zebrafish fell back to g:Profiler. The enrichment step maps these organisms to the Bioconductor OrgDbs org.Sc.sgd.db, org.At.tair.db, org.Ce.eg.db, and org.Dr.eg.db, but those packages were not installed, so the native clusterProfiler GO route (GO over-representation, GO GSEA, and disease ontology) was skipped and the run quietly used the g:Profiler over-representation fallback instead. The four OrgDbs are now in the full environment and the lock, restoring the full GO route for these organisms.

Upgrading

If you already have a bulkseq environment, open Check Environment → Install full R/DESeq2 stack (or re-run it) so it gains r-ashr and the four OrgDbs. On Linux, update the environment from workflow/envs/bulkseq_full.yaml.

Downloads

Windows:

• BulkSeqStudio-Setup-0.12.3.exe — installer
• BulkSeqStudio-Portable-0.12.3.zip — unzip and run BulkSeqStudio.exe, no install

Linux (x86-64, glibc 2.39+):

• BulkSeqStudio-0.12.3-x86_64.AppImage — mark executable and run
• BulkSeqStudio-linux-0.12.3.tar.gz — extract and run BulkSeqStudio

BulkSeq Studio 0.12.2

BulkSeq Studio 0.12.2

Superseded by 0.13.0. A rule guard added in this release has an unescaped-brace bug that Snakemake parses as a format field, aborting the Salmon and HISAT2 routes with a NameError even when the environment is correct. Use 0.13.0 or newer for those aligners. The STAR route and the environment fixes below are unaffected.

Patch release. The Salmon and HISAT2 aligner routes now work with the core environment.

Fixed

• Salmon and HISAT2 aligner routes failed on a core-only environment. The bulkseq_core.yaml profile installed by Install / repair core environment did not include gffread, salmon, or hisat2; those tools were only in the full R/DESeq2 profile. Choosing the Salmon or HISAT2 aligner with a core (or pre-0.11.0) environment ran through trimming and QC, then failed mid-run with exit status 127 (command not found) at make_transcriptome, salmon_index, or hisat2_index. The three tools are now part of the core profile, so every aligner route works with the core environment.
• Check Environment did not probe the alternative-aligner tools. gffread, salmon, and hisat2 are now probed and shown, so a stale environment is visible instead of failing only at run time. The "core ready" indicator still tracks the default STAR route, so a working STAR setup is not reported as incomplete.
• Clearer failure when an aligner tool is missing. The affected rules now exit with a message that points to Setup, instead of a raw exit status 127 partway through the run.

Upgrading

If you already have a bulkseq environment, open Check Environment → Install / repair core environment so it gains gffread, salmon, and hisat2 (an additive update). On Linux, recreate or update the environment from workflow/envs/bulkseq_core.yaml.

Downloads (Windows)

• BulkSeqStudio-Setup-0.12.2.exe — installer
• BulkSeqStudio-Portable-0.12.2.zip — unzip and run BulkSeqStudio.exe, no install