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1.
Zhonghua Kou Qiang Yi Xue Za Zhi ; 56(4): 380-384, 2021 Apr 09.
Artículo en Chino | MEDLINE | ID: mdl-33832042

RESUMEN

Treponema denticola (Td) is a gram-negative anaerobic bacterium closely related to the occurrence and development of periodontal disease and it accounts for a considerable proportion of mature plaque. As a later colonizer of the subgingival plaque biofilm, Td may have complex interactions with earlier and concurrent colonists including symbiotic relationship as while as synergistic or antagonistic effects under the regulation of quorum sensing molecules. Adhesin and coaggregation, mediated by a series of surface molecules, are the basis of the interaction. These interactions are ultimately manifested as gene expression changes in metabolism and virulence, in which are mainly metabolism changes with up- or down-regulation of multiple enzymes related to amino acid metabolism. This article reviews the related researches on the interaction between Td and microorganisms of subgingival plaque.


Asunto(s)
Porphyromonas gingivalis , Treponema denticola , Composición de Base , Filogenia , ARN Ribosómico 16S , Análisis de Secuencia de ADN
2.
Science ; 372(6537): 91-94, 2021 04 02.
Artículo en Inglés | MEDLINE | ID: mdl-33795458

RESUMEN

Neurons are the longest-lived cells in our bodies and lack DNA replication, which makes them reliant on a limited repertoire of DNA repair mechanisms to maintain genome fidelity. These repair mechanisms decline with age, but we have limited knowledge of how genome instability emerges and what strategies neurons and other long-lived cells may have evolved to protect their genomes over the human life span. A targeted sequencing approach in human embryonic stem cell-induced neurons shows that, in neurons, DNA repair is enriched at well-defined hotspots that protect essential genes. These hotspots are enriched with histone H2A isoforms and RNA binding proteins and are associated with evolutionarily conserved elements of the human genome. These findings provide a basis for understanding genome integrity as it relates to aging and disease in the nervous system.


Asunto(s)
Reparación del ADN , Genoma Humano , Inestabilidad Genómica , Neuronas/metabolismo , Envejecimiento/genética , Daño del ADN , ADN Intergénico , Desoxiuridina/análogos & derivados , Desoxiuridina/metabolismo , Células Madre Embrionarias , Histonas/metabolismo , Humanos , Mitosis , Mutación , Enfermedades del Sistema Nervioso/genética , Neuronas/citología , Regiones Promotoras Genéticas , Proteínas de Unión al ARN/metabolismo , Análisis de Secuencia de ADN , Transcripción Genética
3.
BMC Bioinformatics ; 22(1): 119, 2021 Mar 12.
Artículo en Inglés | MEDLINE | ID: mdl-33706720

RESUMEN

BACKGROUND: Metagenomics is the study of microbial genomes for pathogen detection and discovery in human clinical, animal, and environmental samples via Next-Generation Sequencing (NGS). Metagenome de novo sequence assembly is a crucial analytical step in which longer contigs, ideally whole chromosomes/genomes, are formed from shorter NGS reads. However, the contigs generated from the de novo assembly are often very fragmented and rarely longer than a few kilo base pairs (kb). Therefore, a time-consuming extension process is routinely performed on the de novo assembled contigs. RESULTS: To facilitate this process, we propose a new tool for metagenome contig extension after de novo assembly. ContigExtender employs a novel recursive extending strategy that explores multiple extending paths to achieve highly accurate longer contigs. We demonstrate that ContigExtender outperforms existing tools in synthetic, animal, and human metagenomics datasets. CONCLUSIONS: A novel software tool ContigExtender has been developed to assist and enhance the performance of metagenome de novo assembly. ContigExtender effectively extends contigs from a variety of sources and can be incorporated in most viral metagenomics analysis pipelines for a wide variety of applications, including pathogen detection and viral discovery.


Asunto(s)
Genoma Viral , Metagenoma , Metagenómica , Programas Informáticos , Algoritmos , Animales , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Análisis de Secuencia de ADN
4.
Nat Commun ; 12(1): 1583, 2021 03 11.
Artículo en Inglés | MEDLINE | ID: mdl-33707421

RESUMEN

Studies of acute myeloid leukemia rely on DNA sequencing and immunophenotyping by flow cytometry as primary tools for disease characterization. However, leukemia tumor heterogeneity complicates integration of DNA variants and immunophenotypes from separate measurements. Here we introduce DAb-seq, a technology for simultaneous capture of DNA genotype and cell surface phenotype from single cells at high throughput, enabling direct profiling of proteogenomic states in tens of thousands of cells. To demonstrate the approach, we analyze the disease of three patients with leukemia over multiple treatment timepoints and disease recurrences. We observe complex genotype-phenotype dynamics that illustrate the subtlety of the disease process and the degree of incongruity between blast cell genotype and phenotype in different clinical scenarios. Our results highlight the importance of combined single-cell DNA and protein measurements to fully characterize the heterogeneity of leukemia.


Asunto(s)
ADN/genética , Estudios de Asociación Genética , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patología , Análisis de la Célula Individual/métodos , Secuencia de Bases , Línea Celular Tumoral , Técnicas de Genotipaje , Humanos , Inmunofenotipificación , Células Jurkat , Análisis de Secuencia de ADN , Tirosina Quinasa 3 Similar a fms/antagonistas & inhibidores
5.
Klin Lab Diagn ; 66(2): 122-128, 2021 Mar 13.
Artículo en Inglés | MEDLINE | ID: mdl-33734647

RESUMEN

Globally, about 70 million people are infected with the hepatitis C virus (HCV), and about 400 thousand people die annually from chronic hepatitis C complications. The management of patients with chronic hepatitis C may require HCV genotyping, since the efficiency of some widely used antiviral drugs strongly depend on the viral genotype and/or subtype. The most prevalent HCV circulating recombinant form, RF1_2k/1b, is misclassified as genotype 2 by many commercial HCV genotyping kits, based on the RT-PCR analysis of the 5' untranslated region of the HCV genome. This leads to inappropriate patient treatment, since the accepted treatment schemes for HCV genotype 2 are ineffective for the RF1_2k/1b. Here we describe a method for detecting the RNA HCV RF1_2k/1b in blood samples by RT-PCR analysis of two regions in HCV genome (5'UTR and NS5b). The method was tested on 240 blood serum samples from HCV infected patients, in which HCV genotype was defined as 2 or mixed (2+1 or 2+3) by the two commercial genotyping kits "OT-Hepatogen-C genotype" ("DNA-Technology", Moscow) and "RealBest RNA HCV-1/2/3" ("Vector- Best ", Novosibirsk). 50 (20.8%) RF1_2k/1b cases were revealed, including three mixed infections: RF1_2k/1b + 1a, RF1_2k/1b + 3a, RF1_2k/1b + 1b. In all cases, the accuracy of HCV typing by the proposed method was confirmed by Sanger sequencing and phylogenetic analysis. The method is easy to implement into clinical practice and may be used in clinical settings equipped for RT-PCR analysis to correctly identify the recombinant variant RF1_2k/1b.


Asunto(s)
Hepacivirus , Hepatitis C , Genotipo , Hepacivirus/genética , Hepatitis C/diagnóstico , Hepatitis C/genética , Humanos , Moscú , Filogenia , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia de ADN , Suero
6.
Nat Commun ; 12(1): 1504, 2021 03 08.
Artículo en Inglés | MEDLINE | ID: mdl-33686085

RESUMEN

Elucidating functionality in non-coding regions is a key challenge in human genomics. It has been shown that intolerance to variation of coding and proximal non-coding sequence is a strong predictor of human disease relevance. Here, we integrate intolerance to variation, functional genomic annotations and primary genomic sequence to build JARVIS: a comprehensive deep learning model to prioritize non-coding regions, outperforming other human lineage-specific scores. Despite being agnostic to evolutionary conservation, JARVIS performs comparably or outperforms conservation-based scores in classifying pathogenic single-nucleotide and structural variants. In constructing JARVIS, we introduce the genome-wide residual variation intolerance score (gwRVIS), applying a sliding-window approach to whole genome sequencing data from 62,784 individuals. gwRVIS distinguishes Mendelian disease genes from more tolerant CCDS regions and highlights ultra-conserved non-coding elements as the most intolerant regions in the human genome. Both JARVIS and gwRVIS capture previously inaccessible human-lineage constraint information and will enhance our understanding of the non-coding genome.


Asunto(s)
Aprendizaje Profundo , Genoma Humano , Genómica , ADN Intergénico , Variación Genética , Humanos , Análisis de Secuencia de ADN , Secuenciación Completa del Genoma
7.
Zootaxa ; 4927(1): zootaxa.4927.1.1, 2021 Feb 11.
Artículo en Inglés | MEDLINE | ID: mdl-33756717

RESUMEN

DNA barcoding based on a fragment of mitochondrial Cytochrome C Oxidase subunit 1 gene (COI) was applied to the two chironomids Clunio balticus Heimbach (690 base pairs) and C. ponticus Michailova (691 base pairs). The two species differed by one deletion in the nucleotide sequence Adenine. However, the 658-nucleotide long sequences of DNA from the mitochondrial Cytochrome C Oxidase subunit 1 gene (COI) of C. balticus and C. ponticus were identical upon comparison. Further, they compared with homologous sequences for C. marinus Holiday and C. tsushimensis Tokunaga from the Barcode of Life (BOLD) database and the results plotted as a weighted graph, where C. tsushimensis, C. marinus and C. balticus C. ponticus formed three almost equidistant groups. From this, we established that the genetic distance between the respective COI sequences of C. balticus and C. ponticus is minimal, indicating a close relationship between the species indicative of recent common origin. However, the comparative analysis between C. tsushimensis, C. marinus, C. balticus and C. ponticus showed a wider divergence in their respective nucleotide sequences. Overall, our results emphasized that the COI region does not work well as a DNA barcode to identify species within the Clunio genus. Either longer sequences or a multifaceted methodological approach, including morphology, cytogenetic and ecology is needed to distinguish some members of Clunio genus.


Asunto(s)
Chironomidae , Animales , Chironomidae/genética , ADN , Código de Barras del ADN Taxonómico , Filogenia , Análisis de Secuencia de ADN
8.
Zootaxa ; 4949(1): zootaxa.4949.1.8, 2021 Mar 24.
Artículo en Inglés | MEDLINE | ID: mdl-33756999

RESUMEN

Six Anatolian and one European populations of the Myrmeleotettix maculatus species group, which contains M. maculatus and M. ethicus species, have been studied by using molecular genetics methods with mitochondrial COI gene. Myrmeleotettix ethicus is an Anatolian endemic species with local distribution whereas M. maculatus is distributed in western Palearctic. The phylogenetic analysis (ML and BI analyses) of the M. maculatus species group in Anatolia reveals that it consistently recovered two well-supported main clades and four different lineages. Molecular time estimates suggest that the diversification of the M. maculatus species group took place between the Late Tortonian (around 8-9 My) and the Middle of Pliocene-Pleistocene (around 4.3 My-present) periods and the current distribution of the genetic diversity has been affected by the uplifting of the Central Anatolian plateau, the termination of the Messinian salinity crisis, and the Quaternary climatic changes.


Asunto(s)
Saltamontes , Animales , ADN Mitocondrial/genética , Saltamontes/genética , Filogenia , Filogeografía , Análisis de Secuencia de ADN
9.
Microbiome ; 9(1): 58, 2021 03 03.
Artículo en Inglés | MEDLINE | ID: mdl-33658077

RESUMEN

BACKGROUND: Microbial eukaryotes are found alongside bacteria and archaea in natural microbial systems, including host-associated microbiomes. While microbial eukaryotes are critical to these communities, they are challenging to study with shotgun sequencing techniques and are therefore often excluded. RESULTS: Here, we present EukDetect, a bioinformatics method to identify eukaryotes in shotgun metagenomic sequencing data. Our approach uses a database of 521,824 universal marker genes from 241 conserved gene families, which we curated from 3713 fungal, protist, non-vertebrate metazoan, and non-streptophyte archaeplastida genomes and transcriptomes. EukDetect has a broad taxonomic coverage of microbial eukaryotes, performs well on low-abundance and closely related species, and is resilient against bacterial contamination in eukaryotic genomes. Using EukDetect, we describe the spatial distribution of eukaryotes along the human gastrointestinal tract, showing that fungi and protists are present in the lumen and mucosa throughout the large intestine. We discover that there is a succession of eukaryotes that colonize the human gut during the first years of life, mirroring patterns of developmental succession observed in gut bacteria. By comparing DNA and RNA sequencing of paired samples from human stool, we find that many eukaryotes continue active transcription after passage through the gut, though some do not, suggesting they are dormant or nonviable. We analyze metagenomic data from the Baltic Sea and find that eukaryotes differ across locations and salinity gradients. Finally, we observe eukaryotes in Arabidopsis leaf samples, many of which are not identifiable from public protein databases. CONCLUSIONS: EukDetect provides an automated and reliable way to characterize eukaryotes in shotgun sequencing datasets from diverse microbiomes. We demonstrate that it enables discoveries that would be missed or clouded by false positives with standard shotgun sequence analysis. EukDetect will greatly advance our understanding of how microbial eukaryotes contribute to microbiomes. Video abstract.


Asunto(s)
Eucariontes/genética , Eucariontes/aislamiento & purificación , Metagenoma/genética , Metagenómica/métodos , Metagenómica/normas , Animales , Eucariontes/clasificación , Humanos , Análisis de Secuencia de ADN
11.
BMC Bioinformatics ; 22(1): 120, 2021 Mar 12.
Artículo en Inglés | MEDLINE | ID: mdl-33711922

RESUMEN

BACKGROUND: Recently, copy number variations (CNV) impacting genes involved in oncogenic pathways have attracted an increasing attention to manage disease susceptibility. CNV is one of the most important somatic aberrations in the genome of tumor cells. Oncogene activation and tumor suppressor gene inactivation are often attributed to copy number gain/amplification or deletion, respectively, in many cancer types and stages. Recent advances in next generation sequencing protocols allow for the addition of unique molecular identifiers (UMI) to each read. Each targeted DNA fragment is labeled with a unique random nucleotide sequence added to sequencing primers. UMI are especially useful for CNV detection by making each DNA molecule in a population of reads distinct. RESULTS: Here, we present molecular Copy Number Alteration (mCNA), a new methodology allowing the detection of copy number changes using UMI. The algorithm is composed of four main steps: the construction of UMI count matrices, the use of control samples to construct a pseudo-reference, the computation of log-ratios, the segmentation and finally the statistical inference of abnormal segmented breaks. We demonstrate the success of mCNA on a dataset of patients suffering from Diffuse Large B-cell Lymphoma and we highlight that mCNA results have a strong correlation with comparative genomic hybridization. CONCLUSION: We provide mCNA, a new approach for CNV detection, freely available at https://gitlab.com/pierrejulien.viailly/mcna/ under MIT license. mCNA can significantly improve detection accuracy of CNV changes by using UMI.


Asunto(s)
Hibridación Genómica Comparativa , Variaciones en el Número de Copia de ADN , Secuenciación de Nucleótidos de Alto Rendimiento , Adulto , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Análisis de Secuencia de ADN
12.
Zhonghua Bing Li Xue Za Zhi ; 50(3): 190-193, 2021 Mar 08.
Artículo en Chino | MEDLINE | ID: mdl-33677880

RESUMEN

Objective: To investigate the subtypes of H3F3A DNA mutation in H3.3 immunohistochemistry (IHC) negative giant cell tumors of bone (GCTB). Methods: IHC expression of G34W mutated protein was evaluated in 181 cases GCTB. In H3.3 IHC negative cases, Sanger DNA sequencing analysis was used to detect the subtypes H3F3A mutations. Results: Overall, 164 (90.61%) cases of GCTB showed nuclear expression of H3.3, and 17 cases were negative. These 17 H3.3 negative cases were subjected to Sanger DNA sequencing analysis; results showed that eight presented rare mutation subtypes occurring at glycine 34 to leucine (G34L, 3/181, 1.66%), glycine 34 to valine (G34V, 3/181, 1.66%) and glycine 34 to arginine (G34R, 2/181, 1.10%), and the other nine cases were wild type (glycine 34, 9/181, 4.97%). Sanger DNA sequencing analysis confirmed the absence of G34W mutation in the H3.3 negative cases. Combining IHC and DNA sequencing analysis increased the detection rate of H3F3A mutation in the GCTB to 95.03%. Conclusions: H3.3 IHC could detect H3F3A G34W mutation in GCTB, but not for other rare mutation and wild types loci.


Asunto(s)
Neoplasias Óseas , Tumor Óseo de Células Gigantes , Neoplasias Óseas/genética , Tumor Óseo de Células Gigantes/diagnóstico , Tumor Óseo de Células Gigantes/genética , Histonas/genética , Humanos , Inmunohistoquímica , Mutación , Análisis de Secuencia de ADN
13.
Fa Yi Xue Za Zhi ; 37(1): 21-25, 2021 Feb.
Artículo en Inglés, Chino | MEDLINE | ID: mdl-33780180

RESUMEN

Abstract: Objective To study the heteroplasmy of the whole mitochondrial genome genotyping result of hair shaft samples using HID Ion GeneStudioTM S5 Sequencing System. Methods The buccal swabs and blood of 8 unrelated individuals, and hair shaft samples from different parts of the same individual were collected. Amplification of whole mitochondrial genome was performed using Precision ID mtDNA Whole Genome Panel. Analysis and detection of whole mitochondrial genome were carried out using the HID Ion GeneStudioTM S5 Sequencing System. Results The mitochondrial DNA sequences in temporal hair shaft samples from 2 individuals showed heteroplasmy, while whole mitochondrial genome genotyping results of buccal swabs, blood, and hair samples from the other 6 unrelated individuals were consistent. A total of 119 base variations were observed from the 8 unrelated individuals. The numbers of variable sites of the individuals were 29, 40, 38, 35, 13, 36, 40 and 35, respectively. Conclusion Sequence polymorphism can be fully understood using HID Ion GeneStudioTM S5 Sequencing system.


Asunto(s)
ADN Mitocondrial , Genoma Mitocondrial , ADN Mitocondrial/genética , Genoma Mitocondrial/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Análisis de Secuencia de ADN
14.
BMC Bioinformatics ; 22(1): 124, 2021 Mar 16.
Artículo en Inglés | MEDLINE | ID: mdl-33726674

RESUMEN

BACKGROUND: The analysis of long reads or the assessment of assembly or target capture data often necessitates running alignments against reference genomes or gene sets. The aligner outputs are often parsed automatically by scripts, but many kinds of analysis can benefit from the understanding that can follow human inspection of individual alignments. Additionally, diagrams are a useful means of communicating assembly results to others. RESULTS: We developed Alvis, a simple command line tool that can generate visualisations for a number of common alignment analysis tasks. Alvis is a fast and portable tool that accepts input in a variety of alignment formats and will output production ready vector images. Additionally, Alvis will highlight potentially chimeric reads or contigs, a common source of misassemblies. CONCLUSION: Alvis diagrams facilitate improved understanding of assembly quality, enable read coverage to be visualised and potential errors to be identified. Additionally, we found that splitting chimeric reads using the output provided by Alvis can improve the contiguity of assemblies, while maintaining correctness.


Asunto(s)
Quimera , Secuenciación de Nucleótidos de Alto Rendimiento , Análisis de Secuencia de ADN , Visualización de Datos , Genoma , Humanos
15.
BMC Bioinformatics ; 22(1): 115, 2021 Mar 09.
Artículo en Inglés | MEDLINE | ID: mdl-33750296

RESUMEN

BACKGROUND: Today an unprecedented amount of genetic sequence data is stored in publicly available repositories. For decades now, mitochondrial DNA (mtDNA) has been the workhorse of genetic studies, and as a result, there is a large volume of mtDNA data available in these repositories for a wide range of species. Indeed, whilst whole genome sequencing is an exciting prospect for the future, for most non-model organisms' classical markers such as mtDNA remain widely used. By compiling existing data from multiple original studies, it is possible to build powerful new datasets capable of exploring many questions in ecology, evolution and conservation biology. One key question that these data can help inform is what happened in a species' demographic past. However, compiling data in this manner is not trivial, there are many complexities associated with data extraction, data quality and data handling. RESULTS: Here we present the mtDNAcombine package, a collection of tools developed to manage some of the major decisions associated with handling multi-study sequence data with a particular focus on preparing sequence data for Bayesian skyline plot demographic reconstructions. CONCLUSIONS: There is now more genetic information available than ever before and large meta-data sets offer great opportunities to explore new and exciting avenues of research. However, compiling multi-study datasets still remains a technically challenging prospect. The mtDNAcombine package provides a pipeline to streamline the process of downloading, curating, and analysing sequence data, guiding the process of compiling data sets from the online database GenBank.


Asunto(s)
ADN Mitocondrial , Bases de Datos de Ácidos Nucleicos , Teorema de Bayes , ADN Mitocondrial/genética , Análisis de Secuencia de ADN
16.
BMC Bioinformatics ; 22(1): 158, 2021 Mar 25.
Artículo en Inglés | MEDLINE | ID: mdl-33765921

RESUMEN

BACKGROUND: Synthetic long reads (SLR) with long-range co-barcoding information are now widely applied in genomics research. Although several tools have been developed for each specific SLR technique, a robust standalone scaffolder with high efficiency is warranted for hybrid genome assembly. RESULTS: In this work, we developed a standalone scaffolding tool, SLR-superscaffolder, to link together contigs in draft assemblies using co-barcoding and paired-end read information. Our top-to-bottom scheme first builds a global scaffold graph based on Jaccard Similarity to determine the order and orientation of contigs, and then locally improves the scaffolds with the aid of paired-end information. We also exploited a screening algorithm to reduce the negative effect of misassembled contigs in the input assembly. We applied SLR-superscaffolder to a human single tube long fragment read sequencing dataset and increased the scaffold NG50 of its corresponding draft assembly 1349 fold. Moreover, benchmarking on different input contigs showed that this approach overall outperformed existing SLR scaffolders, providing longer contiguity and fewer misassemblies, especially for short contigs assembled by next-generation sequencing data. The open-source code of SLR-superscaffolder is available at https://github.com/BGI-Qingdao/SLR-superscaffolder . CONCLUSIONS: SLR-superscaffolder can dramatically improve the contiguity of a draft assembly by integrating a hybrid assembly strategy.


Asunto(s)
Secuenciación de Nucleótidos de Alto Rendimiento , Programas Informáticos , Algoritmos , Genómica , Humanos , Análisis de Secuencia de ADN
17.
Klin Monbl Augenheilkd ; 238(3): 261-266, 2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-33784789

RESUMEN

Over the past decade, novel high-throughput DNA sequencing technologies have revolutionised both research and diagnostic testing for monogenic disorders. This applies particularly to genetically very heterogeneous disorders like retinal dystrophies (RDs). Next-generation sequencing (NGS) today is considered as reliable as Sanger sequencing, which had been the gold standard for decades. Today, comprehensive NGS-based diagnostic testing reveals the causative mutations in the majority of RD patients, with important implications for genetic counselling for recurrence risks and personalised medical management (from interdisciplinary surveillance to prophylactic measures and, albeit yet rare, [gene] therapy). While DNA sequencing is - in most cases - no longer the diagnostic bottleneck, one needs to be aware of interpretation pitfalls and dead ends. The advent of new (NGS) technologies will solve some of these issues. However, specialised medical geneticists who are familiar with the peculiarities of certain RD genes and closely interact with ophthalmologists will remain key to successful RD research and diagnostic testing for the benefit of the patients. This review sheds light on the current state of the field, its challenges and potential solutions.


Asunto(s)
Distrofias Retinianas , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Mutación/genética , Distrofias Retinianas/diagnóstico , Distrofias Retinianas/genética , Análisis de Secuencia de ADN
18.
Methods Mol Biol ; 2278: 31-44, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33649946

RESUMEN

Genome assembly and annotation are two of the key actions that must be undertaken in order to explore the genomic repertoire of (bifido)bacteria. The gathered information can be employed to genomically characterize a given microorganism, and can also be used to perform comparative genome analysis by including other sequenced (bifido)bacterial strains. Here, we highlight various bioinformatic programs able to manage next generation sequencing data starting from the assembly of a genome to the comparative analyses between strains.


Asunto(s)
Bifidobacterium/genética , Genoma Bacteriano , Genómica/métodos , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Anotación de Secuencia Molecular/métodos , Filogenia , Análisis de Secuencia de ADN/métodos , Secuenciación Completa del Genoma/métodos
19.
Methods Mol Biol ; 2278: 225-232, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33649960

RESUMEN

In this chapter, we present a generic method to achieve de novo genome assembly and methylome analysis of Bifidobacterium genomes using the Pacbio SMRT sequencing and SMRT Link pipeline. The methods described here cover the de novo Pacbio or hybrid Illumina-Pacbio assembly ideally intended to obtain a complete genome sequence, followed by the detection of base modifications and methylome analysis.The identified DNA motifs obtained by methylome analysis can be used to predict active restriction-modification (RM) systems in Bifidobacterium strains. The presence of active RM systems and knowledge on their target motifs may guide selection of suitable cloning or mutagenesis vectors for bifidobacteria.


Asunto(s)
Bifidobacterium/genética , Genoma Bacteriano , Análisis de Secuencia de ADN/métodos , Metilación de ADN , ADN Bacteriano/genética , Motivos de Nucleótidos , Programas Informáticos
20.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 38(3): 282-285, 2021 Mar 10.
Artículo en Chino | MEDLINE | ID: mdl-33751543

RESUMEN

OBJECTIVE: To delineate the characteristics of a novel HLA-DQB1 allele identified during routine HLA matching in a leukemia family. METHODS: The mother and brother of the patient were subjected to PCR sequence-specific oligonucleotide probe (SSOP), PCR sequence-based typ1ing (SBT), as well as next-generation sequencing (NGS). RESULTS: PCR-SBT revealed that the patient's mother and brother's HLA-DQB1 sequences did not fully match with any known allele combination. NGS revealed that the novel allele has differed from the closest matched DQB1*03:02 with a T>G substitution at position 233 in exon 2, which resulted in substitution of Valine at codon 46 by Glycine. Pedigree analysis confirmed that the novel HLA-DQB1 allele was inherited from his mother. CONCLUSION: A novel HLA-DQB1 allele has been identified through next generation sequencing and was officially named as HLA-DQB1*03:362 by the World Health Organization HLA Factor Nomenclature Committee.


Asunto(s)
Cadenas beta de HLA-DQ , Nucleótidos , Análisis de Secuencia de ADN , Alelos , Secuencia de Bases , Cadenas beta de HLA-DQ/genética , Humanos , Masculino , Polimorfismo de Nucleótido Simple
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