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Tuesday, April 21, 2020

The role of genomic structural variation in the genetic improvement of polyploid crops

Many of our major crop species are polyploids, containing more than one genome or set of chromosomes. Polyploid crops present unique challenges, including difficulties in genome assembly, in discriminating between multiple gene and sequence copies, and in genetic mapping, hindering use of genomic data for genetics and breeding. Polyploid genomes may also be more prone to containing structural variation, such as loss of gene copies or sequences (presence–absence variation) and the presence of genes or sequences in multiple copies (copy-number variation). Although the two main types of genomic structural variation commonly identified are presence–absence variation and copy-number variation, we propose…

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Tuesday, April 21, 2020

Improvement of the Pacific bluefin tuna (Thunnus orientalis) reference genome and development of male-specific DNA markers.

The Pacific bluefin tuna, Thunnus orientalis, is a highly migratory species that is widely distributed in the North Pacific Ocean. Like other marine species, T. orientalis has no external sexual dimorphism; thus, identifying sex-specific variants from whole genome sequence data is a useful approach to develop an effective sex identification method. Here, we report an improved draft genome of T. orientalis and male-specific DNA markers. Combining PacBio long reads and Illumina short reads sufficiently improved genome assembly, with a 38-fold increase in scaffold contiguity (to 444 scaffolds) compared to the first published draft genome. Through analysing re-sequence data of 15…

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Tuesday, April 21, 2020

The Single-molecule long-read sequencing of Scylla paramamosain.

Scylla paramamosain is an important aquaculture crab, which has great economical and nutritional value. To the best of our knowledge, few full-length crab transcriptomes are available. In this study, a library composed of 12 different tissues including gill, hepatopancreas, muscle, cerebral ganglion, eyestalk, thoracic ganglia, intestine, heart, testis, ovary, sperm reservoir, and hemocyte was constructed and sequenced using Pacific Biosciences single-molecule real-time (SMRT) long-read sequencing technology. A total of 284803 full-length non-chimeric reads were obtained, from which 79005 high-quality unique transcripts were obtained after error correction and sequence clustering and redundant. Additionally, a total of 52544 transcripts were annotated against…

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Tuesday, April 21, 2020

Large Enriched Fragment Targeted Sequencing (LEFT-SEQ) Applied to Capture of Wolbachia Genomes.

Symbiosis is a major force of evolutionary change, influencing virtually all aspects of biology, from population ecology and evolution to genomics and molecular/biochemical mechanisms of development and reproduction. A remarkable example is Wolbachia endobacteria, present in some parasitic nematodes and many arthropod species. Acquisition of genomic data from diverse Wolbachia clades will aid in the elucidation of the different symbiotic mechanisms(s). However, challenges of de novo assembly of Wolbachia genomes include the presence in the sample of host DNA: nematode/vertebrate or insect. We designed biotinylated probes to capture large fragments of Wolbachia DNA for sequencing using PacBio technology (LEFT-SEQ: Large…

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Tuesday, April 21, 2020

Whole genome sequence and de novo assembly revealed genomic architecture of Indian Mithun (Bos frontalis).

Mithun (Bos frontalis), also called gayal, is an endangered bovine species, under the tribe bovini with 2n?=?58 XX chromosome complements and reared under the tropical rain forests region of India, China, Myanmar, Bhutan and Bangladesh. However, the origin of this species is still disputed and information on its genomic architecture is scanty so far. We trust that availability of its whole genome sequence data and assembly will greatly solve this problem and help to generate many information including phylogenetic status of mithun. Recently, the first genome assembly of gayal, mithun of Chinese origin, was published. However, an improved reference genome…

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Tuesday, April 21, 2020

Long-Read Sequencing Emerging in Medical Genetics

The wide implementation of next-generation sequencing (NGS) technologies has revolutionized the field of medical genetics. However, the short read lengths of currently used sequencing approaches pose a limitation for identification of structural variants, sequencing repetitive regions, phasing alleles and distinguishing highly homologous genomic regions. These limitations may significantly contribute to the diagnostic gap in patients with genetic disorders who have undergone standard NGS, like whole exome or even genome sequencing. Now, the emerging long-read sequencing (LRS) technologies may offer improvements in the characterization of genetic variation and regions that are difficult to assess with the currently prevailing NGS approaches. LRS…

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Tuesday, April 21, 2020

Magic-BLAST, an accurate RNA-seq aligner for long and short reads.

Next-generation sequencing technologies can produce tens of millions of reads, often paired-end, from transcripts or genomes. But few programs can align RNA on the genome and accurately discover introns, especially with long reads. We introduce Magic-BLAST, a new aligner based on ideas from the Magic pipeline.Magic-BLAST uses innovative techniques that include the optimization of a spliced alignment score and selective masking during seed selection. We evaluate the performance of Magic-BLAST to accurately map short or long sequences and its ability to discover introns on real RNA-seq data sets from PacBio, Roche and Illumina runs, and on six benchmarks, and compare…

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Tuesday, April 21, 2020

Improving the sensitivity of long read overlap detection using grouped short k-mer matches.

Single-molecule, real-time sequencing (SMRT) developed by Pacific BioSciences produces longer reads than second-generation sequencing technologies such as Illumina. The increased read length enables PacBio sequencing to close gaps in genome assembly, reveal structural variations, and characterize the intra-species variations. It also holds the promise to decipher the community structure in complex microbial communities because long reads help metagenomic assembly. One key step in genome assembly using long reads is to quickly identify reads forming overlaps. Because PacBio data has higher sequencing error rate and lower coverage than popular short read sequencing technologies (such as Illumina), efficient detection of true overlaps…

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Tuesday, April 21, 2020

Assignment of virus and antimicrobial resistance genes to microbial hosts in a complex microbial community by combined long-read assembly and proximity ligation.

We describe a method that adds long-read sequencing to a mix of technologies used to assemble a highly complex cattle rumen microbial community, and provide a comparison to short read-based methods. Long-read alignments and Hi-C linkage between contigs support the identification of 188 novel virus-host associations and the determination of phage life cycle states in the rumen microbial community. The long-read assembly also identifies 94 antimicrobial resistance genes, compared to only seven alleles in the short-read assembly. We demonstrate novel techniques that work synergistically to improve characterization of biological features in a highly complex rumen microbial community.

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Tuesday, April 21, 2020

Long-read based de novo assembly of low-complexity metagenome samples results in finished genomes and reveals insights into strain diversity and an active phage system.

Complete and contiguous genome assemblies greatly improve the quality of subsequent systems-wide functional profiling studies and the ability to gain novel biological insights. While a de novo genome assembly of an isolated bacterial strain is in most cases straightforward, more informative data about co-existing bacteria as well as synergistic and antagonistic effects can be obtained from a direct analysis of microbial communities. However, the complexity of metagenomic samples represents a major challenge. While third generation sequencing technologies have been suggested to enable finished metagenome-assembled genomes, to our knowledge, the complete genome assembly of all dominant strains in a microbiome sample…

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Tuesday, April 21, 2020

A comparative evaluation of hybrid error correction methods for error-prone long reads.

Third-generation sequencing technologies have advanced the progress of the biological research by generating reads that are substantially longer than second-generation sequencing technologies. However, their notorious high error rate impedes straightforward data analysis and limits their application. A handful of error correction methods for these error-prone long reads have been developed to date. The output data quality is very important for downstream analysis, whereas computing resources could limit the utility of some computing-intense tools. There is a lack of standardized assessments for these long-read error-correction methods.Here, we present a comparative performance assessment of ten state-of-the-art error-correction methods for long reads. We…

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