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…
Interested to learn about pangenomes? Explore this guide to learn how they provide a more complete picture of the core genes of a given species and how that can provide better biological understanding.
Many scientists are using PacBio Single Molecule, Real-Time (SMRT) Sequencing to explore the genomes and transcriptomes of a wide variety of marine species and ecosystems. These studies are already adding to our understanding of how marine species adapt and evolve, contributing to conservation efforts, and informing how we can optimize food production through efficient aquaculture.
Despite apparent carbon limitation, anoxic deep subsurface brines at the Soudan Underground Iron Mine harbor active microbial communities. To characterize these assemblages, we performed shotgun metagenomics of native and enriched samples. Following enrichment on poised electrodes and long read sequencing, we recovered from the metagenome the closed, circular genome of a novel Desulfuromonas sp. with remarkable genomic features that were not fully resolved by short read assembly alone. This organism was essentially absent in unenriched Soudan communities, indicating that electrodes are highly selective for putative metal reducers. Native community metagenomes suggest that carbon cycling is driven by methyl-C1 metabolism, in…
Animals in the phylum Hemichordata have provided key understanding of the origins and development of body patterning and nervous system organization. However, efforts to sequence and assemble the genomes of highly heterozygous non-model organisms have proven to be difficult with traditional short read approaches. Long repetitive DNA structures, extensive structural variation between haplotypes in polyploid species, and large genome sizes are limiting factors to achieving highly contiguous genome assemblies. Here we present the highly contiguous de novo assembly and preliminary annotation of an indirect developing hemichordate genome, Schizocardium californicum, using SMRT Sequening long reads.
In this webinar, Barbara Block of Stanford University and Paul Peluso of PacBio describe how plant and animal whole genome sequencing remains a challenging endeavor, particularly due to genome size, high density of repetitive elements, and heterozygosity. Because of this, often only a single, fragmented reference genome is available for a species, genus, or even family, limiting the ability to answer important biological questions. Looking at the trends in genome assembly and annotation over the past year, such as pan-genomes and phasing, this webinar explores how Single Molecule, Real-Time (SMRT) Sequencing is utilized to develop long-lasting genomic resources, supporting research…
Microbial Assembly is our latest pipeline, specifically designed to assemble bacterial genomes (between 2 and 10 Mb) and plasmids. This pipeline includes the implementation of a new, circular-aware read alignment tool (Raptor), among other algorithmic improvements, which will be covered in this webinar. The topics covered include, staged assembly of bacterial chromosomes and plasmids, implementation of Raptor, a circular-aware read aligner, himeric read detection, origin of replication orientation, troubleshooting and more.
The widespread occurrence of repetitive stretches of DNA in genomes of organisms across the tree of life imposes fundamental challenges for sequencing, genome assembly, and automated annotation of genes and proteins. This multi-level problem can lead to errors in genome and protein databases that are often not recognized or acknowledged. As a consequence, end users working with sequences with repetitive regions are faced with ‘ready-to-use’ deposited data whose trustworthiness is difficult to determine, let alone to quantify. Here, we provide a review of the problems associated with tandem repeat sequences that originate from different stages during the sequencing-assembly-annotation-deposition workflow, and…
Antarcticibacterium flavum JB01H24T was isolated from a marine sediment of the Ross Sea, Antarctica. Whole-genome sequencing of the strain Antarcticibacterium flavum JB01H24T was achieved using PacBio RS II platform. The resulting complete genome comprised of one closed, complete chromosome of 4,319,074 base pairs with a 40.87% G?+?C content, where genomic analyses demonstrated that it is constituted mostly by putative ORFs with unknown functions, representing a novel genetic feature. It is the first complete genome sequence of the Antarcticibacterium strain.
Although many ß-agarases that hydrolyze the ß-1,4 linkages of agarose have been biochemically characterized, only three a-agarases that hydrolyze the a-1,3 linkages are reported to date. In this study, a new a-agarase, AgaWS5, from Catenovulum sediminis WS1-A, a new agar-degrading marine bacterium, was biochemically characterized. AgaWS5 belongs to the glycoside hydrolase (GH) 96 family. AgaWS5 consists of 1295 amino acids (140 kDa) and has the 65% identity to an a-agarase, AgaA33, obtained from an agar-degrading bacterium Thalassomonas agarivorans JAMB-A33. AgaWS5 showed the maximum activity at a pH and temperature of 8 and 40 °C, respectively. AgaWS5 showed a cold-tolerance, and…
Large yellow croaker (Larimichthys crocea) is an economically important marine fish species of China. Due to overfishing and marine pollution, the wild stocks of this croaker have collapsed in the past decades. Meanwhile, the cultured croaker is facing the difficulties of reduced genetic diversity and low growth rate. To explore the molecular markers related to the growth traits of croaker and providing the related SNPs for the marker-assisted selection, we used double-digest restriction-site associated DNA (ddRAD) sequencing to dissect the genetic bases of growth traits in a cultured population and identify the SNPs that associated with important growth traits by…
Marinobacter sp. strain LQ44, an alkaliphile and moderate halophile from a deep-sea hydrothermal vent on the East Pacific Rise, is a novel phenol-degrading bacterium that is capable of utilizing phenol as sole carbon and energy sources. Here, we present the complete genome sequence of strain LQ44, which consists of 4,435,564?bp with a circular chromosome, 4164 protein-coding genes, 3 rRNA operons and 50 tRNAs. Genome analysis revealed that strain LQ44 may degrade phenol via meta-cleavage pathway. The LQ44 genome contains multiple genes involved in pH adaptation and osmotic adjustment. Genes related to hydrocarbon degradation, aerobic denitrification and potential industrial important enzymes…
The zebra mussel, Dreissena polymorpha, continues to spread from its native range in Eurasia to Europe and North America, causing billions of dollars in damage and dramatically altering invaded aquatic ecosystems. Despite these impacts, there are few genomic resources for Dreissena or related bivalves, with nearly 450 million years of divergence between zebra mussels and its closest sequenced relative. Although the D. polymorpha genome is highly repetitive, we have used a combination of long-read sequencing and Hi-C-based scaffolding to generate the highest quality molluscan assembly to date. Through comparative analysis and transcriptomics experiments we have gained insights into processes that…
Paracoccus sp. Arc7-R13, a silver nanoparticles (AgNPs) synthesizing bacterium, was isolated from Arctic Ocean sediment. Here we describe the complete genome of Paracoccus sp. Arc7-R13. The complete genome contains 4,040,012?bp with 66.66?mol%?G?+?C content, including one circular chromosome of 3,231,929?bp (67.45?mol%?G?+?C content), and eight plasmids with length ranging from 24,536?bp to 199,685?bp. The genome contains 3835 protein-coding genes (CDSs), 49 tRNA genes, as well as 3 rRNA operons as 16S-23S-5S rRNA. Based on the gene annotation and Swiss-Prot analysis, a total of 15 genes belonging to 11 kinds, including silver exporting P-type ATPase (SilP), alkaline phosphatase, nitroreductase, thioredoxin reductase, NADPH dehydrogenase…
Two Marinobacter sp. NP-4 and NP-6 were isolated from a deep oceanic basaltic crust at North Pond, located at the western flank of the Mid-Atlantic Ridge. These two strains are capable of using multiple carbon sources such as acetate, succinate, glucose and sucrose while take oxygen as a primary electron acceptor. The strain NP-4 is also able to grow anaerobically under 20?MPa, with nitrate as the electron acceptor, thus represents a piezotolerant. To explore the metabolic potentials of Marinobacter sp. NP-4 and NP-6, the complete genome of NP-4 and close-to-complete genome of NP-6 were sequenced. The genome of NP-4 contains…