A Rust reimplementation of STAR (Spliced Transcripts Alignment to a Reference), the widely-used RNA-seq aligner originally written in C++ by Alexander Dobin.

Overview

rustar-aligner aims to be a faithful port of STAR, matching the original behavior as closely as possible. It uses the same genome index format, accepts the same --camelCase command-line parameters, and produces compatible SAM/BAM output.

Current status: End-to-end single-end and paired-end RNA-seq alignment with splice junction detection, two-pass mode, chimeric alignment detection (including multi-junction Tier 3), gene-level quantification, and multi-threaded parallel processing. 396 tests passing (383 unit + 8 integration + others), 0 clippy warnings.

Quick Start

Build

cargo build --release

Generate genome index

target/release/rustar-aligner --runMode genomeGenerate \
  --genomeDir /path/to/genome_index \
  --genomeFastaFiles /path/to/genome.fa

Align reads

target/release/rustar-aligner \
  --genomeDir /path/to/genome_index \
  --readFilesIn reads.fq \
  --outSAMtype SAM \
  --outSAMstrandField intronMotif \
  --outFileNamePrefix /path/to/output_

Paired-end alignment

target/release/rustar-aligner \
  --genomeDir /path/to/genome_index \
  --readFilesIn reads_1.fq reads_2.fq \
  --outSAMtype SAM \
  --outFileNamePrefix /path/to/output_

BAM output

target/release/rustar-aligner \
  --genomeDir /path/to/genome_index \
  --readFilesIn reads.fq \
  --outSAMtype BAM Unsorted \
  --outFileNamePrefix /path/to/output_

Coordinate-sorted BAM output

target/release/rustar-aligner \
  --genomeDir /path/to/genome_index \
  --readFilesIn reads.fq \
  --outSAMtype BAM SortedByCoordinate \
  --outFileNamePrefix /path/to/output_

Two-pass mode

target/release/rustar-aligner \
  --genomeDir /path/to/genome_index \
  --readFilesIn reads.fq \
  --twopassMode Basic \
  --outFileNamePrefix /path/to/output_

Gene-level counts

target/release/rustar-aligner \
  --genomeDir /path/to/genome_index \
  --readFilesIn reads.fq \
  --sjdbGTFfile /path/to/annotation.gtf \
  --quantMode GeneCounts \
  --outFileNamePrefix /path/to/output_

Accuracy Comparison vs STAR

Benchmarked on 10,000 yeast RNA-seq reads (150 bp, ERR12389696), compared to STAR 2.7.x with identical parameters and genome index.

Single-End (10k reads, 150 bp SE)

Metric	rustar-aligner	STAR
Unique mapped	82.6%	82.6%
Multi-mapped	7.4%	7.4%
Total mapped	90.0%	90.0%
Position agreement	96.5% raw / 99.815% tie-adjusted	—
STAR-only reads	0	—
rustar-aligner-only reads	0	—
CIGAR-only diffs	1 (seed-level tie in homopolymer)	—

Tie-adjusted: 299 of 313 disagreements are verified genuine ties — both tools find identical alignment sets but select different copies due to SA-order or RNG tie-breaking differences. Excluding these, faithfulness is 99.815% (8,611/8,627 non-tie reads exact).

Paired-End (10k read pairs, 150 bp)

Metric	rustar-aligner	STAR
Both mates mapped	8,390	8,390
Half-mapped pairs	0	0
Unmapped pairs	0	0
PE faithfulness (tie-adjusted)	99.883%	—
MAPQ inflations	0	—
MAPQ deflations	0	—
NH tag diffs	0	—
Proper-pair diffs	0	—

PE faithfulness: 16,284 / 16,306 mate alignments exactly match STAR (same position, CIGAR, MAPQ, proper-pair flag, and NH tag). 475 diffs excluded as tie-breaking differences (same MAPQ+NH, different repeat copy chosen).

Supported Features

• Single-end and paired-end alignment with mate rescue
• SAM, unsorted BAM, and coordinate-sorted BAM output (--outSAMtype SAM, BAM Unsorted, or BAM SortedByCoordinate)
• Multi-threaded parallel alignment (--runThreadN)
• GTF-based junction annotation with scoring bonus (--sjdbGTFfile)
• Two-pass mode for novel junction discovery (--twopassMode Basic)
• SJDB insertion into genome index at genomeGenerate time
• Chimeric alignment detection — SE and PE, 4-tier pipeline: transcript-pair search, multi-cluster, soft-clip re-seeding, residual outer re-seeding for multi-junction fusions (--chimSegmentMin)
• Gene-level read counting (--quantMode GeneCounts → ReadsPerGene.out.tab)
• Transcriptome-coordinate SAM output (--quantMode TranscriptomeSAM)
• Post-alignment read filtering (--outFilterType BySJout)
• Splice junction output (SJ.out.tab)
• Unmapped read output to FASTQ (--outReadsUnmapped Fastx → Unmapped.out.mate1 / mate2)
• Gzip-compressed FASTQ input (--readFilesCommand zcat)
• Read group tags (--outSAMattrRGline)
• Seeded RNG for reproducible tie-breaking (--runRNGseed)
• SAM optional tags: NH, HI, AS, NM, nM, XS, jM, jI, MD
• --outSAMattributes control (Standard/All/None/explicit list)
• SECONDARY flag (0x100) on multi-mapper alignments
• Configurable output limits (--outSAMmultNmax)
• Bidirectional seed search with scoreSeedBest pre-extension
• Junction boundary optimization (jR scanning)
• Log.final.out statistics file (STAR-compatible, MultiQC-parseable)

Known Limitations

• No STARsolo single-cell features (Phase 14, deferred)

See ROADMAP.md for detailed implementation tracking.

Building from Source

Requires Rust 2024 edition (rustc 1.85+).

cargo build --release    # Release build
cargo test               # Run tests
cargo clippy             # Lint
cargo fmt                # Format

Development

The majority of rustar-aligner's code was written by Claude Code (Anthropic's AI coding assistant), with technical direction, architecture decisions, and validation by the project maintainer.

License

MIT (matching the original STAR license)