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Thursday, November 12, 2020

Technical Note: Preparing samples for PacBio whole genome sequencing for de novo assembly – Collection and storage

Single Molecule, Real-Time (SMRT) Sequencing uses the natural process of DNA replication to sequence long fragments of native DNA. As such, starting with high-quality, high molecular weight (HMW) genomic DNA (gDNA) will result in better sequencing performance across difficult to sequence regions of the genome. To obtain the highest quality, long DNA it is important to start with sample types compatible with HMW DNA extraction methods. This technical note is intended to give general guidance on sample collection, preparation, and storage across a range of commonly encountered sample types used for SMRT Sequencing whole genome projects. It is important to…

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Tuesday, April 21, 2020

Identification and characterization of chicken circovirus from commercial broiler chickens in China.

Circoviruses are found in many species, including mammals, birds, lower vertebrates and invertebrates. To date, there are no reports of circovirus-induced diseases in chickens. In this study, we identified a new strain of chicken circovirus (CCV) by PacBio third-generation sequencing samples from chickens with acute gastroenteritis in a Shandong commercial broiler farm in China. The complete genome of CCV was verified by inverse PCR. Genomic analysis revealed that CCV codes two inverse open reading frames (ORFs), and a potential stem-loop structure was present at the 5′ end with a structure typical of a circular virus. Phylogenetic tree analysis showed that…

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Tuesday, April 21, 2020

A draft nuclear-genome assembly of the acoel flatworm Praesagittifera naikaiensis.

Acoels are primitive bilaterians with very simple soft bodies, in which many organs, including the gut, are not developed. They provide platforms for studying molecular and developmental mechanisms involved in the formation of the basic bilaterian body plan, whole-body regeneration, and symbiosis with photosynthetic microalgae. Because genomic information is essential for future research on acoel biology, we sequenced and assembled the nuclear genome of an acoel, Praesagittifera naikaiensis.To avoid sequence contamination derived from symbiotic microalgae, DNA was extracted from embryos that were free of algae. More than 290x sequencing coverage was achieved using a combination of Illumina (paired-end and mate-pair…

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Tuesday, April 21, 2020

Mycobacterium ulcerans Population Genomics To Inform on the Spread of Buruli Ulcer across Central Africa.

Buruli ulcer is a neglected tropical disease of skin and subcutaneous tissue caused by infection with the pathogen Mycobacterium ulcerans Many critical issues for disease control, such as understanding the mode of transmission and identifying source reservoirs of M. ulcerans, are still largely unknown. Here, we used genomics to reconstruct in detail the evolutionary trajectory and dynamics of M. ulcerans populations at a central African scale and at smaller geographical village scales. Whole-genome sequencing (WGS) data were analyzed from 179 M. ulcerans strains isolated from all Buruli ulcer foci in the Democratic Republic of the Congo, The Republic of Congo,…

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Tuesday, April 21, 2020

Expedited assessment of terrestrial arthropod diversity by coupling Malaise traps with DNA barcoding 1.

Monitoring changes in terrestrial arthropod communities over space and time requires a dramatic increase in the speed and accuracy of processing samples that cannot be achieved with morphological approaches. The combination of DNA barcoding and Malaise traps allows expedited, comprehensive inventories of species abundance whose cost will rapidly decline as high-throughput sequencing technologies advance. Aside from detailing protocols from specimen sorting to data release, this paper describes their use in a survey of arthropod diversity in a national park that examined 21?194 specimens representing 2255 species. These protocols can support arthropod monitoring programs at regional, national, and continental scales.

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Tuesday, April 21, 2020

Complete genome sequence of Streptomyces spongiicola HNM0071T, a marine sponge-associated actinomycete producing staurosporine and echinomycin

Streptomyes spongiicola HNM0071T is a novel marine sponge-associated actinomycete with potential to produce antitumor agents including staurosporine and echinomycin. Here, we present the complete genome sequence of S. spongiicola HNM0071, which consists of a linear chromosome of 7,180,417?bp, 5669 protein coding genes, 18 rRNA genes, and 66 tRNA genes. Twenty-seven putative secondary metabolite biosynthetic gene clusters were found in the genome. Among them, the staurosporine and echinomycin gene clusters have been described completely. The complete genome information presented here will enable us to investigate the biosynthetic mechanism of two well-known antitumor antibiotics and to discover novel secondary metabolites with potential…

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Tuesday, April 21, 2020

Do the toll-like receptors and complement systems play equally important roles in freshwater adapted Dolly Varden char (Salvelinus malma)?

Unlike the normal anadromous lifestyle, Chinese native Dolly Varden char (Salvelinus malma) is locked in land and lives in fresh water lifetime. To explore the effect of freshwater adaption on its immune system, we constructed a pooled cDNA library of hepatopancreas and spleen of Chinese freshwater Dolly Varden char (S. malma). A total of 27,829 unigenes were generated from 31,233 high-quality transcripts and 17,670 complete open reading frames (ORF) were identified. Totally 25,809 unigenes were successfully annotated and it classified more native than adaptive immunity-associated genes, and more genes involved in toll-like receptor signal pathway than those in complement and…

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Tuesday, April 21, 2020

Complete genome of Pseudoalteromonas atlantica ECSMB14104, a Gammaproteobacterium inducing mussel settlement

Pseudoalteromonas is widely distributed in the marine environments and the biofilms formed by Pseudoalteromonas promote settlement of many species of invertebrates. Here, we show the complete genome of Pseudoalteromonas atlantica ECSMB14104, which was isolated from biofilms formed in the East China Sea and exhibited inducing activity on the Mytilus coruscus settlement. Complete genome of this strain containsa total of 3325 genes and the GC content of 41.02%. This genomic information is contributed to molecular mechanism of P. atlantica ECSMB14104 regulating mussel settlement.

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Tuesday, April 21, 2020

A full-length transcriptome of Sepia esculenta using a combination of single-molecule long-read (SMRT) and Illumina sequencing

As an economically important cephalopods species, wild-caught Sepia esculenta fishery has suffered a server decline due to over-fishing and ocean environmental damage. To restore this seriously declining fishery resource, we should understand the genetic foundation and molecular mechanism of spawning, reproduction and mortal of golden cuttlefish. In this study, we generated the full-length transcriptome of S. esculenta based on the total RNA of tissue samples (brain, optic gland, nidamental gland, ovary and muscle at different developmental stages) using a combination of single-molecule real-time (SMRT) and Illumina RNA-seq technology. A total of 14.16 Gb SMRT sequencing data were assembled into 94,635…

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Tuesday, April 21, 2020

Full-length transcriptome analysis of Litopenaeus vannamei reveals transcript variants involved in the innate immune system.

To better understand the immune system of shrimp, this study combined PacBio isoform sequencing (Iso-Seq) and Illumina paired-end short reads sequencing methods to discover full-length immune-related molecules of the Pacific white shrimp, Litopenaeus vannamei. A total of 72,648 nonredundant full-length transcripts (unigenes) were generated with an average length of 2545 bp from five main tissues, including the hepatopancreas, cardiac stomach, heart, muscle, and pyloric stomach. These unigenes exhibited a high annotation rate (62,164, 85.57%) when compared against NR, NT, Swiss-Prot, Pfam, GO, KEGG and COG databases. A total of 7544 putative long noncoding RNAs (lncRNAs) were detected and 1164 nonredundant…

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Tuesday, April 21, 2020

Full-length transcriptome sequences obtained by a combination of sequencing platforms applied to heat shock proteins and polyunsaturated fatty acids biosynthesis in Pyropia haitanensis

Pyropia haitanensis is a high-yield commercial seaweed in China. Pyropia haitanensis farms often suffer from problems such as severe germplasm degeneration, while the mechanisms underlying resistance to abiotic stresses remain unknown because of lacking genomic information. Although many previous studies focused on using next-generation sequencing (NGS) technologies, the short-read sequences generated by NGS generally prevent the assembly of full-length transcripts, and then limit screening functional genes. In the present study, which was based on hybrid sequencing (NGS and single-molecular real-time sequencing) of the P. haitanensis thallus transcriptome, we obtained high-quality full-length transcripts with a mean length of 2998 bp and…

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Tuesday, April 21, 2020

SMRT long reads and Direct Label and Stain optical maps allow the generation of a high-quality genome assembly for the European barn swallow (Hirundo rustica rustica).

The barn swallow (Hirundo rustica) is a migratory bird that has been the focus of a large number of ecological, behavioral, and genetic studies. To facilitate further population genetics and genomic studies, we present a reference genome assembly for the European subspecies (H. r. rustica).As part of the Genome10K effort on generating high-quality vertebrate genomes (Vertebrate Genomes Project), we have assembled a highly contiguous genome assembly using single molecule real-time (SMRT) DNA sequencing and several Bionano optical map technologies. We compared and integrated optical maps derived from both the Nick, Label, Repair, and Stain technology and from the Direct Label…

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Tuesday, April 21, 2020

Genetic basis of functional variability in adhesion G protein-coupled receptors.

The enormous sizes of adhesion G protein-coupled receptors (aGPCRs) go along with complex genomic exon-intron architectures giving rise to multiple mRNA variants. There is a need for a comprehensive catalog of aGPCR variants for proper evaluation of the complex functions of aGPCRs found in structural, in vitro and animal model studies. We used an established bioinformatics pipeline to extract, quantify and visualize mRNA variants of aGPCRs from deeply sequenced transcriptomes. Data analysis showed that aGPCRs have multiple transcription start sites even within introns and that tissue-specific splicing is frequent. On average, 19 significantly expressed transcript variants are derived from a…

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