Genomes and metagenomes · What you can do here
Assemble a genome from sequencing reads
Assemble genomes de novo using MaSuRCA from short paired-end reads alone or combined with long reads, then evaluate assembly contiguity and lineage-specific completeness with BUSCO.
What this involves
De novo genome assembly reconstructs contiguous chromosomes from sequencing reads without a reference sequence. MaSuRCA takes Illumina paired-end reads and extends them into super-reads. For hybrid assemblies incorporating Oxford Nanopore or PacBio CLR data, these super-reads correct long reads into mega-reads before assembly using the CABOG engine. Every MaSuRCA run requires Illumina paired-end data to provide baseline accuracy and error correction.
Selecting the right data combination determines the assembly's final contiguity. While short reads alone assemble bacteria and eukaryotes into many fragmented contigs because reads cannot span repeats longer than the library insert size, hybrid assemblies using long reads can yield complete bacterial chromosomes in a single scaffold. For hybrid workflows, MaSuRCA requires at least 50x Illumina paired-end coverage and consumes up to 25x of the longest available reads.
Assembly quality requires distinct checks on contiguity and gene completeness. While MaSuRCA outputs primary contigs, scaffolds, and alternative assemblies for heterozygous loci, contiguity metrics such as N50 do not measure gene content. Completeness must be independently evaluated with BUSCO against conserved orthologue sets, separating biological duplication from uncollapsed haplotypes.
How it goes
- 1Input preparation and sizingProvide untrimmed Illumina paired-end reads alongside Oxford Nanopore or PacBio CLR files or accession numbers, specifying the estimated genome size in megabases to configure hashing and bacterial settings.
- 2De novo assembly with MaSuRCAMaSuRCA 4.1.4 generates super-reads, constructs mega-reads from any long reads provided, and executes assembly via CABOG on a dedicated machine.
- 3Consensus polishingFor hybrid runs, MaSuRCA applies its POLCA tool to polish consensus sequence substitutions and indels using the short reads, producing primary.polished.fasta.
- 4Completeness evaluation with BUSCOBUSCO 6.1.0 evaluates the polished or primary assembly in genome mode against a chosen OrthoDB lineage set, quantifying complete, duplicated, fragmented, and missing orthologues.
Start it here
Ask in plain language. The agent works on a Linux machine of its own — it installs and runs what the job needs, keeps every file, and reads the results back with the numbers explained. Nothing to set up, and a long run continues after you close the page.
What to watch for
- Do not trim, clean, or correct reads before running MaSuRCA, as the assembler performs its own internal error correction and expects raw reads.
- Do not input PacBio HiFi reads or attempt long-read-only assemblies into MaSuRCA; it is strictly designed for short reads alone or short reads combined with PacBio CLR or Nanopore reads.
- Assemblies projected to exceed the platform's 24-hour job ceiling, such as 500 Mb or 1 Gb genomes, should use the generated config.txt to run directly on an external cluster rather than starting an execution that cannot complete.
- Contig N50, derived from breaking scaffolds at runs of Ns, is the reliable measure of sequence contiguity because scaffold N50 includes artificial gap spans.
- High duplication in BUSCO results may indicate uncollapsed haplotypes or sample contamination rather than true whole-genome duplication.
- Scores from BUSCO are only comparable when evaluated against the exact same lineage set and OrthoDB generation.
What it runs
Programs
- MaSuRCARuns as a job on its own machine· GPL-3.0Assembles genomes from short Illumina reads, alone or combined with long reads, using super-reads.
- BUSCORuns as a job on its own machine· Software MIT; lineage datasets CC BY-ND 4.0Assesses completeness of genome assemblies, gene sets, and transcriptomes using universal single-copy orthologs.
- fastpPreinstalled on the research machineQuality control and preprocessing software that trims, filters and profiles short-read FASTQ sequencing data in one pass.
- SeqKitPreinstalled on the research machineA cross-platform toolkit for fast inspection, filtering, conversion, and analysis of FASTA and FASTQ files.
nf-core pipelines
Questions
Do I need to install anything to assemble a genome from sequencing reads?
No. The research agent does this work on a Linux machine of its own — it runs MaSuRCA, BUSCO, fastp, SeqKit there, keeps the files, and reads the results back to you. Long runs continue if you close the page.
Should I filter or quality-trim my reads before starting MaSuRCA?
No. The authors of MaSuRCA explicitly instruct users not to trim, clean, or correct sequencing reads prior to assembly, as the assembler handles its own read correction.
Can I assemble a genome using only Oxford Nanopore reads in MaSuRCA?
No. Every MaSuRCA assembly requires Illumina paired-end reads to construct super-reads; Nanopore reads are only used as an addition to Illumina data for hybrid assembly.
Which output file contains the final assembled sequence?
For hybrid assemblies, the primary output file to use is primary.polished.fasta produced by POLCA. For Illumina-only runs, use primary.genome.scf.fasta. The alternative.genome.scf.fasta file contains redundant scaffolds and alternative haplotypes set aside by the assembler.
What is the difference between contig N50 and scaffold N50?
Scaffold N50 measures the length of assembled sequences including unresolved gap regions filled with Ns, whereas contig N50 breaks scaffolds across those gap runs to report actual contiguous sequence lengths.
Written from the method the research agent follows for this work and the documentation of the tools listed above. Statements citing a figure that does not appear there are removed before publication. Corollary is a non-commercial research demonstration; results are not for clinical use.