Two high-quality de novo genomes from single ethanol-preserved specimens of tiny metazoans (Collembola)

Abstract

Background Genome sequencing of all known eukaryotes on Earth promises unprecedented advances in biological sciences and in biodiversity-related applied fields such as environmental management and natural product research. Advances in long-read DNA sequencing make it feasible to generate high-quality genomes for many non–genetic model species. However, long-read sequencing today relies on sizable quantities of high-quality, high molecular weight DNA, which is mostly obtained from fresh tissues. This is a challenge for biodiversity genomics of most metazoan species, which are tiny and need to be preserved immediately after collection. Here we present de novo genomes of 2 species of submillimeter Collembola. For each, we prepared the sequencing library from high molecular weight DNA extracted from a single specimen and using a novel ultra-low input protocol from Pacific Biosciences. This protocol requires a DNA input of only 5 ng, permitted by a whole-genome amplification step. Results The 2 assembled genomes have N50 values >5.5 and 8.5 Mb, respectively, and both contain ∼96% of BUSCO genes. Thus, they are highly contiguous and complete. The genomes are supported by an integrative taxonomy approach including placement in a genome-based phylogeny of Collembola and designation of a neotype for 1 of the species. Higher heterozygosity values are recorded in the more mobile species. Both species are devoid of the biosynthetic pathway for β-lactam antibiotics known in several Collembola, confirming the tight correlation of antibiotic synthesis with the species way of life. Conclusions It is now possible to generate high-quality genomes from single specimens of minute, field-preserved metazoans, exceeding the minimum contig N50 (1 Mb) required by the Earth BioGenome Project.

Highly contiguous assemblies of 101 drosophilid genomes

“…Many other species are simply understudied and sample availability is limited to a few flies collected from the wild and possibly preserved in ethanol for many years. Methods for assembling genomes with small quantities of DNA from single insects ( Adams et al, 2020 ; Kingan et al, 2019 ; Schneider et al, 2021 ) or dealing with degraded specimens from older collections will be particularly important as the scope of future work expands beyond stock center and laboratory lines.…”

The USDA-ARS Ag100Pest Initiative: High-Quality Genome Assemblies for Agricultural Pest Arthropod Research

“…Our continuous integration and refinement of new methods to address particular challenges posed by arthropods have allowed Ag100Pest to sequence species that were not tractable when we began this project. Specifically, the reduction in input DNA requirements since the project’s inception has generated low and ultra-low input protocols for long-read sequencing libraries [ 66 , 70 ] that have allowed us to sequence species with very small physical sizes. Additionally, PacBio’s optimization of circular consensus sequencing (CCS) greatly increased the sequencing accuracy and generation of High-Fidelity (HiFi) reads [ 33 ], which hold many benefits over CLR.…”

Mabs, a suite of tools for gene-informed genome assembly

“…In principle, this strategy of “gene-informed parameter optimization” may be applied to any genome assembler that has some parameters that affect its algorithm. I have chosen Hifiasm and Flye because in multiple benchmarks, they were shown to be the best or among the best assemblers for accurate (PacBio HiFi) and error-prone (Oxford Nanopore Technologies and PacBio CLR) reads, respectively (Jung et al , 2020; Murigneux et al , 2020; Dida and Yi, 2021; Gavrielatos et al , 2021; Guiglielmoni et al , 2021; Schneider et al , 2021; Wick and Holt, 2021; Rabanal et al , 2022; Xie et al , 2022; Zhang et al , 2022). Since Hifiasm and Flye were favourably compared with other genome assemblers many times, in this work, I compare Mabs only with Hifiasm and Flye and do not make comparisons with other assemblers.…”