Ultralow Input Genome Sequencing Library Preparation from a Single Tardigrade Specimen

Yoshida, Yuki; Konno, Sayuri; Nishino, Ryousuke; Murai, Yumi; Tomita, Masaru; Arakawa, Kazuharu

doi:10.3791/57615

Cited by 9 publications

(6 citation statements)

References 19 publications

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“…The steady development of sequencing technologies is allowing the generation of genomes spanning the diversity of life, which now includes minute organisms. Indeed, thanks to the latest low and ultra-low DNA input protocols, sequencing high-quality genomes from millimetric animals is now possible ( Yoshida et al, 2018 ; Schneider et al, 2021 ; Lord et al, 2023 ). In this study, we used the Pacbio Ultra-Low DNA Input protocol to sequence the genome of N. westbladi , reporting the first nemertodermatid genome, sequenced from a single microscopic worm.…”

Section: Discussionmentioning

confidence: 99%

The draft genome of the microscopic Nemertoderma westbladi sheds light on the evolution of Acoelomorpha genomes

Abalde,

Tellgren-Roth,

Heintz

et al. 2023

Front. Genet.

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Background: Xenacoelomorpha is a marine clade of microscopic worms that is an important model system for understanding the evolution of key bilaterian novelties, such as the excretory system. Nevertheless, Xenacoelomorpha genomics has been restricted to a few species that either can be cultured in the lab or are centimetres long. Thus far, no genomes are available for Nemertodermatida, one of the group’s main clades and whose origin has been dated more than 400 million years ago.Methods: DNA was extracted from a single specimen and sequenced with HiFi following the PacBio Ultra-Low DNA Input protocol. After genome assembly, decontamination, and annotation, the genome quality was benchmarked using two acoel genomes and one Illumina genome as reference. The gene content of three cnidarians, three acoelomorphs, four deuterostomes, and eight protostomes was clustered in orthogroups to make inferences of gene content evolution. Finally, we focused on the genes related to the ultrafiltration excretory system to compare patterns of presence/absence and gene architecture among these clades.Results: We present the first nemertodermatid genome sequenced from a single specimen of Nemertoderma westbladi. Although genome contiguity remains challenging (N50: 60 kb), it is very complete (BUSCO: 80.2%, Metazoa; 88.6%, Eukaryota) and the quality of the annotation allows fine-detail analyses of genome evolution. Acoelomorph genomes seem to be relatively conserved in terms of the percentage of repeats, number of genes, number of exons per gene and intron size. In addition, a high fraction of genes present in both protostomes and deuterostomes are absent in Acoelomorpha. Interestingly, we show that all genes related to the excretory system are present in Xenacoelomorpha except Osr, a key element in the development of these organs and whose acquisition seems to be interconnected with the origin of the specialised excretory system.Conclusion: Overall, these analyses highlight the potential of the Ultra-Low Input DNA protocol and HiFi to generate high-quality genomes from single animals, even for relatively large genomes, making it a feasible option for sequencing challenging taxa, which will be an exciting resource for comparative genomics analyses.

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Section: Discussionmentioning

confidence: 99%

The draft genome of the microscopic Nemertoderma westbladi sheds light on the evolution of Acoelomorpha genomes

Abalde,

Tellgren-Roth,

Heintz

et al. 2023

Front. Genet.

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“…On the other hand, no sustainable culture of heterotardigrades has been established to date. Therefore, we employed an ultra-low-input genome sequencing protocol that we developed previously [38][39][40] to achieve whole genome sequencing from a single specimen of tardigrade containing less than 50 pg of DNA collected from the wild. However, such a sample requires strong precautions to minimize the possibility of contamination 23,38,42 .…”

Section: Discussionmentioning

confidence: 99%

“…We used the ultra-low-input library sequencing protocol established in our previous study 39,40 . In brief, genomic DNA was extracted using a Quick-gDNA Microprep Kit (Zymo Research) after lysis with two freeze-thaw cycles of -80°C and 37°C incubation, and the extracted DNA was sheared to 550-bp target fragments with Covaris M220.…”

Section: Genome and Transcriptome Sequencingmentioning

confidence: 99%

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Multiomics study of a heterotardigrade,Echinisicus testudo, suggests convergent evolution of anhydrobiosis-related proteins in Tardigrada

Murai

Yagi-Utsumi

Tomita

et al. 2020

Preprint

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Many limno-terrestrial tardigrades can enter an ametabolic state upon desiccation, in which the animals can withstand extreme environments. To date, studies of the molecular mechanism have predominantly investigated the class Eutardigrada, and that in the Heterotardigrada, remains elusive. To this end, we report a multiomics study of the heterotardigrade Echiniscus testudo, which is one of the most desiccation-tolerant species. None of the previously identified tardigrade-specific anhydrobiosis-related genes was conserved, while the loss and expansion of existing pathways were partly shared. Furthermore, we identified two families of novel abundant heat-soluble proteins and the proteins exhibited structural changes from random coil to α-helix as the water content decreased in vitro. These characteristics are analogous to those of anhydrobiosis-related protein in eutardigrades, while there is no conservation at the sequence level. Our results suggest that Heterotardigrada have partly shared but distinct anhydrobiosis machinery compared with Eutardigrada, possibly due to convergent evolution within Tardigrada.

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“…Studies that used H. exemplaris strain Z151 prior to the 2018 species disambiguation from H. dujardini (Gasiorek et al 2018) generally referred to the species as H. dujardini, or as Hypsibius cf. dujardini, and in many of these studies the strain was indicated as Z151 and/or sourced from Sciento Bavan et al 2009;Mali et al 2010;Cesari et al 2012;Beltrán-Pardo et al 2013;Horikawa et al 2013;Tenlen et al 2013;Smith and Jockusch 2014;Boothby et al 2015Boothby et al , 2017Gross and Mayer 2015;Kondo et al 2015;Arakawa et al 2016;Bemm et al 2016;Fernandez et al 2016;Hering et al 2016;Hyra et al 2016a,b;Kosztyła et al 2016;Koutsovoulos et al 2016;Levin et al 2016;Smith et al 2016Stec et al 2016;Erdmann et al 2017;Gross et al 2017Gross et al , 2018Vasanthan et al 2017;Yoshida et al 2017Yoshida et al , 2018Fadero et al 2018;Nelson 2018;Rost-Roszkowska et al 2018). One study using Thulinia stephaniae (Hejnol and Schnabel 2005) indicates some contrasting developmental features in a second tardigrade species.…”

Section: Related Speciesmentioning

confidence: 99%

The Emergence of the TardigradeHypsibius exemplarisas a Model System

Goldstein

2018

Cold Spring Harb Protoc

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The success of scientists in revealing biological mechanisms has depended in large part on choosing tractable model systems. In 1997, molecular phylogenetics revealed that two of biology's most tractable models-Caenorhabditis elegans and Drosophila-are much more closely related to each other than had been thought previously. I began to explore whether any of the little-studied members of this branch of the tree of life might serve as a new model for comparative biology that could make use of the rich and ongoing sources of information flowing from C. elegans and Drosophila research. Tardigrades, also known as water bears, make up a phylum of microscopic animals. The tardigrade Hypsibius exemplaris (recently disambiguated from a closely related species, Hypsibius dujardini) can be maintained in laboratories and has a generation time of <2 wk at room temperature. Stocks of animals can be stored frozen and revived. The animals and their embryos are optically clear, and embryos are laid in groups, with each synchronous clutch of embryos laid in a clear molt. We have developed techniques for laboratory study of this system, including methods for microinjection of animals, immunolocalization, in situ hybridization, RNA interference, transcriptomics, and methods for identifying proteins that mediate tolerance to extreme environments. Here, I review the development of this animal as an emerging model system, as well as recent molecular studies aimed at understanding the evolution of developmental mechanisms that underpin the evolution of animal form and at understanding how biological materials can survive extreme environments.

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Ultralow Input Genome Sequencing Library Preparation from a Single Tardigrade Specimen

Cited by 9 publications

References 19 publications

The draft genome of the microscopic Nemertoderma westbladi sheds light on the evolution of Acoelomorpha genomes

The draft genome of the microscopic Nemertoderma westbladi sheds light on the evolution of Acoelomorpha genomes

Multiomics study of a heterotardigrade,Echinisicus testudo, suggests convergent evolution of anhydrobiosis-related proteins in Tardigrada

The Emergence of the TardigradeHypsibius exemplarisas a Model System

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