Long- read sequencing and de novo assembly of the cynomolgus macaque genome

AbstractLong-read de novo genome assembly continues to advance rapidly. However, there is a lack of effective tools to accurately evaluate the assembly results, especially for structural errors. We present Inspector, a reference-free long-read de novo assembly evaluator which faithfully reports types of errors and their precise locations. Notably, Inspector can correct the assembly errors based on consensus sequences derived from raw reads covering erroneous regions. Based on in silico and long-read assembly results from multiple long-read data and assemblers, we demonstrate that in addition to providing generic metrics, Inspector can accurately identify both large-scale and small-scale assembly errors.

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Oxford Nanopore Sequencing, Hybrid Error Correction, and de novo Assembly of a Eukaryotic Genome

10.1101/013490 ◽

2015 ◽

Cited By ~ 23

Author(s):

Sara Goodwin ◽

James Gurtowski ◽

Scott Ethe-Sayers ◽

Panchajanya Deshpande ◽

Michael Schatz ◽

...

Keyword(s):

Error Correction ◽

De Novo Assembly ◽

De Novo ◽

Correction Algorithm ◽

Membrane Pore ◽

Complete Representation ◽

Oxford Nanopore ◽

Long Read ◽

Error Correction Algorithm ◽

Sequencing Instrument

Monitoring the progress of DNA molecules through a membrane pore has been postulated as a method for sequencing DNA for several decades. Recently, a nanopore-based sequencing instrument, the Oxford Nanopore MinION, has become available that we used for sequencing the S. cerevisiae genome. To make use of these data, we developed a novel open-source hybrid error correction algorithm Nanocorr (https://github.com/jgurtowski/nanocorr) specifically for Oxford Nanopore reads, as existing packages were incapable of assembling the long read lengths (5-50kbp) at such high error rate (between ~5 and 40% error). With this new method we were able to perform a hybrid error correction of the nanopore reads using complementary MiSeq data and produce a de novo assembly that is highly contiguous and accurate: the contig N50 length is more than ten-times greater than an Illumina-only assembly (678kb versus 59.9kbp), and has greater than 99.88% consensus identity when compared to the reference. Furthermore, the assembly with the long nanopore reads presents a much more complete representation of the features of the genome and correctly assembles gene cassettes, rRNAs, transposable elements, and other genomic features that were almost entirely absent in the Illumina-only assembly.

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De novo assembly of a Tibetan genome and identification of novel structural variants associated with high-altitude adaptation

National Science Review ◽

10.1093/nsr/nwz160 ◽

2019 ◽

Vol 7 (2) ◽

pp. 391-402 ◽

Cited By ~ 3

Author(s):

Yaoxi He ◽

Haiyi Lou ◽

Chaoying Cui ◽

Lian Deng ◽

Yang Gao ◽

...

Keyword(s):

High Altitude ◽

De Novo Assembly ◽

De Novo ◽

Population Analysis ◽

Extreme Environments ◽

Structural Variants ◽

Base Pairs ◽

High Quality ◽

Long Read ◽

Altitude Adaptation

Abstract Structural variants (SVs) may play important roles in human adaptation to extreme environments such as high altitude but have been under-investigated. Here, combining long-read sequencing with multiple scaffolding techniques, we assembled a high-quality Tibetan genome (ZF1), with a contig N50 length of 24.57 mega-base pairs (Mb) and a scaffold N50 length of 58.80 Mb. The ZF1 assembly filled 80 remaining N-gaps (0.25 Mb in total length) in the reference human genome (GRCh38). Markedly, we detected 17 900 SVs, among which the ZF1-specific SVs are enriched in GTPase activity that is required for activation of the hypoxic pathway. Further population analysis uncovered a 163-bp intronic deletion in the MKL1 gene showing large divergence between highland Tibetans and lowland Han Chinese. This deletion is significantly associated with lower systolic pulmonary arterial pressure, one of the key adaptive physiological traits in Tibetans. Moreover, with the use of the high-quality de novo assembly, we observed a much higher rate of genome-wide archaic hominid (Altai Neanderthal and Denisovan) shared non-reference sequences in ZF1 (1.32%–1.53%) compared to other East Asian genomes (0.70%–0.98%), reflecting a unique genomic composition of Tibetans. One such archaic hominid shared sequence—a 662-bp intronic insertion in the SCUBE2 gene—is enriched and associated with better lung function (the FEV1/FVC ratio) in Tibetans. Collectively, we generated the first high-resolution Tibetan reference genome, and the identified SVs may serve as valuable resources for future evolutionary and medical studies.

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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

BMC Microbiology ◽

10.1186/s12866-019-1500-0 ◽

2019 ◽

Vol 19 (1) ◽

Cited By ~ 31

Author(s):

Vincent Somerville ◽

Stefanie Lutz ◽

Michael Schmid ◽

Daniel Frei ◽

Aline Moser ◽

...

Keyword(s):

De Novo Assembly ◽

De Novo ◽

Low Complexity ◽

Strain Diversity ◽

Long Read

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De novo assembly of Dekkera bruxellensis: a multi technology approach using short and long-read sequencing and optical mapping

GigaScience ◽

10.1186/s13742-015-0094-1 ◽

2015 ◽

Vol 4 (1) ◽

Cited By ~ 17

Author(s):

Remi-Andre Olsen ◽

Ignas Bunikis ◽

Ievgeniia Tiukova ◽

Kicki Holmberg ◽

Britta Lötstedt ◽

...

Keyword(s):

De Novo Assembly ◽

De Novo ◽

Optical Mapping ◽

Dekkera Bruxellensis ◽

Long Read

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Advancing the genetic utility of pre-clinical species through a high-quality assembly of the cynomolgus monkey (Macaca fascicularis) genome

10.1101/2020.05.01.072280 ◽

2020 ◽

Author(s):

Elias Oziolor ◽

Shawn Sullivan ◽

Hayley Mangelson ◽

Stephen M. Eacker ◽

Michael Agostino ◽

...

Keyword(s):

Biomedical Research ◽

Genetic Architecture ◽

Cynomolgus Macaque ◽

Hybrid Assembly ◽

Primate Model ◽

Sequencing Technologies ◽

Macaque Genome ◽

Long Read ◽

Demographic Estimation ◽

Human Primate

AbstractThe cynomolgus macaque is a non-human primate model, heavily used in biomedical research, but with outdated genomic resources. Here we have used the latest long-read sequencing technologies in order to assemble a fully phased, chromosome-level assembly for the cynomolgus macaque. We have built a hybrid assembly with PacBio, 10x Genomics, and HiC technologies, resulting in a diploid assembly that spans a length of 5.1 Gb with a total of 16,741 contigs (N50 of 0.86Mb) contained in 370 scaffolds (N50 of 138 Mb) positioned on 42 chromosomes (21 homologous pairs). This assembly is highly homologous to former assemblies and identifies novel inversions and provides higher confidence in the genetic architecture of the cynomolgus macaque genome. A demographic estimation is also able to capture the recent genetic bottleneck in the Mauritius population, from which the sequenced individual originates. We offer this resource as an enablement for genetic tools to be built around this important model for biomedical research.

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De novo diploid genome assembly for genome-wide structural variant detection

NAR Genomics and Bioinformatics ◽

10.1093/nargab/lqz018 ◽

2019 ◽

Vol 2 (1) ◽

Author(s):

Lu Zhang ◽

Xin Zhou ◽

Ziming Weng ◽

Arend Sidow

Keyword(s):

De Novo Assembly ◽

De Novo ◽

Pairwise Alignment ◽

Cost Effective ◽

Difficult Problem ◽

Ancestral State ◽

Fundamental Limitations ◽

Human Genomes ◽

Genome Wide ◽

Long Read

Abstract Detection of structural variants (SVs) on the basis of read alignment to a reference genome remains a difficult problem. De novo assembly, traditionally used to generate reference genomes, offers an alternative for SV detection. However, it has not been applied broadly to human genomes because of fundamental limitations of short-fragment approaches and high cost of long-read technologies. We here show that 10× linked-read sequencing supports accurate SV detection. We examined variants in six de novo 10× assemblies with diverse experimental parameters from two commonly used human cell lines: NA12878 and NA24385. The assemblies are effective for detecting mid-size SVs, which were discovered by simple pairwise alignment of the assemblies’ contigs to the reference (hg38). Our study also shows that the base-pair level SV breakpoint accuracy is high, with a majority of SVs having precisely correct sizes and breakpoints. Setting the ancestral state of SV loci by comparing to ape orthologs allows inference of the actual molecular mechanism (insertion or deletion) causing the mutation. In about half of cases, the mechanism is the opposite of the reference-based call. We uncover 214 SVs that may have been maintained as polymorphisms in the human lineage since before our divergence from chimp. Overall, we show that de novo assembly of 10× linked-read data can achieve cost-effective SV detection for personal genomes.

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