LongStitch: High-quality genome assembly correction and scaffolding using long reads

Abstract

Background Generating high-quality de novo genome assemblies is foundational to the genomics study of model and non-model organisms. In recent years, long-read sequencing has greatly benefited genome assembly and scaffolding, a process by which assembled sequences are ordered and oriented through the use of long-range information. Long reads are better able to span repetitive genomic regions compared to short reads, and thus have tremendous utility for resolving problematic regions and helping generate more complete draft assemblies. Here, we present LongStitch, a scalable pipeline that corrects and scaffolds draft genome assemblies exclusively using long reads. Results LongStitch incorporates multiple tools developed by our group and runs in up to three stages, which includes initial assembly correction (Tigmint-long), followed by two incremental scaffolding stages (ntLink and ARKS-long). Tigmint-long and ARKS-long are misassembly correction and scaffolding utilities, respectively, previously developed for linked reads, that we adapted for long reads. Here, we describe the LongStitch pipeline and introduce our new long-read scaffolder, ntLink, which utilizes lightweight minimizer mappings to join contigs. LongStitch was tested on short and long-read assemblies of three different human individuals using corresponding nanopore long-read data, and improves the contiguity of each assembly from 2.0-fold up to 304.6-fold (as measured by NGA50 length). Furthermore, LongStitch generates more contiguous and correct assemblies compared to state-of-the-art long-read scaffolder LRScaf in most tests, and consistently runs in under five hours using less than 23GB of RAM. Conclusions Due to its effectiveness and efficiency in improving draft assemblies using long reads, we expect LongStitch to benefit a wide variety of de novo genome assembly projects. The LongStitch pipeline is freely available at https://github.com/bcgsc/longstitch.

Post-embryonic tail development through molting of the freshwater shrimpNeocaridina denticulata

“…The five assemblies were combined into a single assembly using quickmerge version 0.3 (44). Subsequently, the assembly was scaffolded three times using ntlink version 1.3.9 and Flye polishing version 2.9.1 (45,46). Haplotype duplication was removed and the mitochondrial genome was eliminated using purge_haplotigs version 1.1.2 (47).…”

High-quality genome of the zoophytophagous stink bug,Nesidiocoris tenuis, informs their food habit adaptation

“…Genome contamination from symbiotic microorganisms was removed by MEGAN and DIAMOND (Bagci et al, 2021). We lastly performed 1-kbp or less raw long-read assembly by using ntLink with the "-gap_fill" option (Coombe et al, 2023).…”

“…Scaffolding was performed with the Hi-C reads by using SALSA v2.3 (Ghurye et al, 2019). We next performed raw longread-based scaffolding by using ntLink (Coombe et al, 2023) and then finally checked manually for remaining gaps. We used GENESPACE (Wang et al, 2012) for the synteny comparison of scaffolded genomes and visualization of the results.…”