Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 10 de 10
Filtrar
Mais filtros










Base de dados
Tipo de estudo
Intervalo de ano de publicação
1.
Nat Commun ; 10(1): 4053, 2019 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-31492841

RESUMO

Whole genome sequencing (WGS) studies have estimated the human germline mutation rate per basepair per generation (~1.2 × 10-8) to be higher than in mice (3.5-5.4 × 10-9). In humans, most germline mutations are paternal in origin and numbers of mutations per offspring increase with paternal and maternal age. Here we estimate germline mutation rates and spectra in six multi-sibling mouse pedigrees and compare to three multi-sibling human pedigrees. In both species we observe a paternal mutation bias, a parental age effect, and a highly mutagenic first cell division contributing to the embryo. We also observe differences between species in mutation spectra, in mutation rates per cell division, and in the parental bias of mutations in early embryogenesis. These differences between species likely result from both species-specific differences in cellular genealogies of the germline, as well as biological differences within the same stage of embryogenesis or gametogenesis.

2.
Genome Res ; 27(10): 1704-1714, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28855261

RESUMO

Structural mosaic abnormalities are large post-zygotic mutations present in a subset of cells and have been implicated in developmental disorders and cancer. Such mutations have been conventionally assessed in clinical diagnostics using cytogenetic or microarray testing. Modern disease studies rely heavily on exome sequencing, yet an adequate method for the detection of structural mosaicism using targeted sequencing data is lacking. Here, we present a method, called MrMosaic, to detect structural mosaic abnormalities using deviations in allele fraction and read coverage from next-generation sequencing data. Whole-exome sequencing (WES) and whole-genome sequencing (WGS) simulations were used to calculate detection performance across a range of mosaic event sizes, types, clonalities, and sequencing depths. The tool was applied to 4911 patients with undiagnosed developmental disorders, and 11 events among nine patients were detected. For eight of these 11 events, mosaicism was observed in saliva but not blood, suggesting that assaying blood alone would miss a large fraction, possibly >50%, of mosaic diagnostic chromosomal rearrangements.


Assuntos
Exoma , Genoma Humano , Mosaicismo , Análise de Sequência de DNA/métodos , Feminino , Humanos , Masculino , Análise de Sequência de DNA/instrumentação
3.
Nat Commun ; 8(1): 303, 2017 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-28827725

RESUMO

Heterozygous mutations within homozygous sequences descended from a recent common ancestor offer a way to ascertain de novo mutations across multiple generations. Using exome sequences from 3222 British-Pakistani individuals with high parental relatedness, we estimate a mutation rate of 1.45 ± 0.05 × 10-8 per base pair per generation in autosomal coding sequence, with a corresponding non-crossover gene conversion rate of 8.75 ± 0.05 × 10-6 per base pair per generation. This is at the lower end of exome mutation rates previously estimated in parent-offspring trios, suggesting that post-zygotic mutations contribute little to the human germ-line mutation rate. We find frequent recurrence of mutations at polymorphic CpG sites, and an increase in C to T mutations in a 5' CCG 3' to 5' CTG 3' context in the Pakistani population compared to Europeans, suggesting that mutational processes have evolved rapidly between human populations.Estimates of human mutation rates differ substantially based on the approach. Here, the authors present a multi-generational estimate from the autozygous segment in a non-European population that gives insight into the contribution of post-zygotic mutations and population-specific mutational processes.


Assuntos
Genética Populacional/métodos , Genoma Humano/genética , Taxa de Mutação , Mutação , Exoma/genética , Mutação em Linhagem Germinativa , Heterozigoto , Homozigoto , Humanos , Polimorfismo Genético
4.
Nature ; 543(7647): 714-718, 2017 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-28329761

RESUMO

Somatic cells acquire mutations throughout the course of an individual's life. Mutations occurring early in embryogenesis are often present in a substantial proportion of, but not all, cells in postnatal humans and thus have particular characteristics and effects. Depending on their location in the genome and the proportion of cells they are present in, these mosaic mutations can cause a wide range of genetic disease syndromes and predispose carriers to cancer. They have a high chance of being transmitted to offspring as de novo germline mutations and, in principle, can provide insights into early human embryonic cell lineages and their contributions to adult tissues. Although it is known that gross chromosomal abnormalities are remarkably common in early human embryos, our understanding of early embryonic somatic mutations is very limited. Here we use whole-genome sequences of normal blood from 241 adults to identify 163 early embryonic mutations. We estimate that approximately three base substitution mutations occur per cell per cell-doubling event in early human embryogenesis and these are mainly attributable to two known mutational signatures. We used the mutations to reconstruct developmental lineages of adult cells and demonstrate that the two daughter cells of many early embryonic cell-doubling events contribute asymmetrically to adult blood at an approximately 2:1 ratio. This study therefore provides insights into the mutation rates, mutational processes and developmental outcomes of cell dynamics that operate during early human embryogenesis.


Assuntos
Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário/genética , Mutação , Adulto , Células Sanguíneas/metabolismo , Linhagem da Célula/genética , Genoma Humano/genética , Mutação em Linhagem Germinativa/genética , Humanos , Mosaicismo , Mutagênese , Taxa de Mutação
5.
Hum Mutat ; 38(4): 390-399, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27995740

RESUMO

Fcγ receptors are a family of cell-surface receptors that are expressed by a host of different innate and adaptive immune cells, and mediate inflammatory responses by binding the Fc portion of immunoglobulin G. In humans, five low-affinity receptors are encoded by the genes FCGR2A, FCGR2B, FCGR2C, FCGR3A, and FCGR3B, which are located in an 82.5-kb segmental tandem duplication on chromosome 1q23.3, which shows extensive copy-number variation (CNV). Deletions of FCGR3B have been suggested to increase the risk of inflammatory diseases such as systemic lupus erythematosus and rheumatoid arthritis (RA). In this study, we identify the deletion breakpoints of FCGR3B deletion alleles in the UK population and endogamous native American population, and show that some but not all alleles are likely to be identical-by-descent. We also localize a duplication breakpoint, confirming that the mechanism of CNV generation is nonallelic homologous recombination, and identify several alleles with gene conversion events using fosmid sequencing data. We use information on the structure of the deletion alleles to distinguish FCGR3B deletions from FCGR3A deletions in whole-genome array comparative genomic hybridization (aCGH) data. Reanalysis of published aCGH data using this approach supports association of FCGR3B deletion with increased risk of RA in a large cohort of 1,982 cases and 3,271 controls (odds ratio 1.61, P = 2.9×10-3 ).


Assuntos
Artrite Reumatoide/genética , Variações do Número de Cópias de DNA , Predisposição Genética para Doença/genética , Receptores de IgG/genética , Deleção de Sequência , Alelos , Artrite Reumatoide/metabolismo , Estudos de Coortes , Hibridização Genômica Comparativa/métodos , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Haplótipos , Recombinação Homóloga , Humanos , Polimorfismo de Nucleotídeo Único , Receptores de IgG/metabolismo , Fatores de Risco
6.
Methods Mol Biol ; 1400: 95-106, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26895048

RESUMO

With the advent of new generations of high-throughput sequencing technologies, the catalog of human genome variants created by retrotransposon activity is expanding rapidly. However, despite these advances in describing L1 diversity and the fact that L1 must retrotranspose in the germline or prior to germline partitioning to be evolutionarily successful, direct assessment of de novo L1 retrotransposition in the germline or early embryogenesis has not been achieved for endogenous L1 elements. A direct study of de novo L1 retrotransposition into susceptible loci within sperm DNA (Freeman et al., Hum Mutat 32(8):978-988, 2011) suggested that the rate of L1 retrotransposition in the germline is much lower than previously estimated (<1 in 400 individuals versus 1 in 9 individuals (Kazazian, Nat Genet 22(2):130, 1999). Based on these revised estimates of the L1 retrotransposition rate, we modified the ATLAS L1 display technique (Badge et al., Am J Hum Genet 72(4):823-838, 2003) to investigate de novo L1 retrotransposition in human genomes. In this chapter, we describe how we combined a high-coverage ATLAS variant with high-throughput sequencing, achieving 11-25× sequence depth per single amplicon, to study L1 retrotransposition in whole genome amplified (WGA) DNAs.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala , Elementos Nucleotídeos Longos e Dispersos , Tipagem Molecular , Reação em Cadeia da Polimerase , Biologia Computacional/métodos , Genoma Humano , Biblioteca Genômica , Genômica/métodos , Humanos , Tipagem Molecular/métodos , Reação em Cadeia da Polimerase/métodos
7.
Nat Genet ; 48(2): 126-133, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26656846

RESUMO

Germline mutations are a driving force behind genome evolution and genetic disease. We investigated genome-wide mutation rates and spectra in multi-sibling families. The mutation rate increased with paternal age in all families, but the number of additional mutations per year differed by more than twofold between families. Meta-analysis of 6,570 mutations showed that germline methylation influences mutation rates. In contrast to somatic mutations, we found remarkable consistency in germline mutation spectra between the sexes and at different paternal ages. In parental germ line, 3.8% of mutations were mosaic, resulting in 1.3% of mutations being shared by siblings. The number of these shared mutations varied significantly between families. Our data suggest that the mutation rate per cell division is higher during both early embryogenesis and differentiation of primordial germ cells but is reduced substantially during post-pubertal spermatogenesis. These findings have important consequences for the recurrence risks of disorders caused by de novo mutations.


Assuntos
Mutação em Linhagem Germinativa , Ilhas de CpG , Feminino , Humanos , Masculino , Mosaicismo , Idade Paterna , Linhagem
8.
Hum Mutat ; 34(7): 974-85, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23553801

RESUMO

Long INterspersed Element-1 (LINE-1 or L1) retrotransposons are the only autonomously active transposable elements in the human genome. The average human genome contains ∼80-100 active L1s, but only a subset of these L1s are highly active or 'hot'. Human L1s are closely related in sequence, making it difficult to decipher progenitor/offspring relationships using traditional phylogenetic methods. However, L1 mRNAs can sometimes bypass their own polyadenylation signal and instead utilize fortuitous polyadenylation signals in 3' flanking genomic DNA. Retrotransposition of the resultant mRNAs then results in lineage specific sequence "tags" (i.e., 3' transductions) that mark the descendants of active L1 progenitors. Here, we developed a method (Transduction-Specific Amplification Typing of L1 Active Subfamilies or TS-ATLAS) that exploits L1 3' transductions to identify active L1 lineages in a genome-wide context. TS-ATLAS enabled the characterization of a putative active progenitor of one L1 lineage that includes the disease causing L1 insertion L1RP , and the identification of new retrotransposition events within two other "hot" L1 lineages. Intriguingly, the analysis of the newly discovered transduction lineage members suggests that L1 polyadenylation, even within a lineage, is highly stochastic. Thus, TS-ATLAS provides a new tool to explore the dynamics of L1 lineage evolution and retrotransposon biology.


Assuntos
Genoma Humano/genética , Elementos Nucleotídeos Longos e Dispersos/genética , Mutagênese Insercional/métodos , Retroelementos/genética , DNA/genética , Humanos , Poliadenilação
9.
Mol Biotechnol ; 47(3): 243-52, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20872285

RESUMO

Intracisternal A-type particle (IAP) elements are high copy number long terminal repeat (LTR) rodent retrotransposons. Some IAP elements can transpose, and are responsible for ~12% of spontaneous mouse mutations. Inbred mouse strains show variation in genomic IAP distribution, contributing to inter-strain genetic variability. Additionally IAP elements can influence the transcriptional regulation of neighbouring genes through their strong LTR promoter, effecting phenotypic variation. This genetic and phenotypic variability can translate into experimental variability between mouse strains. For example, it has been demonstrated that strain-specific genetic/epigenetic factors can interact to yield variable responses to drugs. Therefore, in experimental contexts it is essential to unequivocally identify mouse strains. Recently it was estimated that any two inbred strains share only ~40% of their IAP insertions. Of the remaining 60%, some insertions will be strain specific, fixed during inbreeding. These fixed insertions can be exploited as genetic markers to identify inbred strains, if they can be identified simply and efficiently. Here, we report the development of a PCR-based system allowing direct acquisition of strain-specific IAP insertions. In a pilot study, we identified 21 IAP loci, genotyped IAP insertions at 9 loci, and discovered two strain-specific insertions that could reliably identify these strains.


Assuntos
Genes de Partícula A Intracisternal/genética , Retroelementos/genética , Sequências Repetidas Terminais/genética , Animais , Genótipo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Endogâmicos DBA , Reação em Cadeia da Polimerase
10.
Biotechniques ; 46(4): 277-84, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19450234

RESUMO

The HeLa cell line is the oldest, most widely distributed, permanent human cell line. As a nearly ubiquitous inhabitant of laboratories using tissue culture techniques, its aggressive growth characteristics make it a problematic contaminant that can overgrow less robust cell lines. Consequently, HeLa contamination is common in both the research laboratory and cell line repository contexts, and its detection is hampered by the lack of a rapid, sensitive and robust assay. Here we report the development of a HeLa-specific DNA diagnostic test: a single duplex detection PCR assay targeting an L1 retrotransposon insertion. All HeLa clones from a geographically diverse panel were positive by this assay, and the particular L1 insertion we identified appears to be unique to the HeLa cell line. The assay can detect very low levels of HeLa contamination (<1%), and can be performed on un-purified cell pellets, allowing rapid routine screening.


Assuntos
Biomarcadores/análise , Células Cultivadas , Células HeLa , Elementos Nucleotídeos Longos e Dispersos/genética , Contaminação de Equipamentos , Frequência do Gene , Humanos , Reação em Cadeia da Polimerase
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA