Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 16 de 16
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Methods Mol Biol ; 2794: 293-304, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38630238

RESUMO

Droplet digital PCR (ddPCR) is an emerging method for the absolute quantification of PCR products, and it can detect DNA copy numbers accurately. It analyzes the end-point absolute fluorescence signals of the PCR-positive droplets and calculates the target concentration. EvaGreen is a nonspecific double-stranded DNA-binding fluorescent dye, and the ddPCR system also supports assays using this cost-effective hydrolysis probe. Here, we describe a simple method of quantification for DNA copy numbers using the EvaGreen single-color fluorescent design.


Assuntos
Variações do Número de Cópias de DNA , Genômica , Corantes Fluorescentes , Reação em Cadeia da Polimerase , DNA/genética
2.
Am J Med Genet A ; 191(6): 1632-1638, 2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-36916329

RESUMO

Many disease-causing genes have been identified by determining the breakpoints of balanced chromosomal translocations. Recent progress in genomic analysis has accelerated the analysis of chromosomal translocation-breakpoints at the nucleotide level. Using a long-read whole-genome sequence, we analyzed the breakpoints of the cytogenetically balanced chromosomal translocation t(5;15)(q21;26.3), which was confirmed to be of de novo origin, in a patient with a neurodevelopmental disorder. The results showed complex rearrangements with seven fragments consisting of five breakpoint-junctions (BJs). Four of the five BJs showed microhomologies of 1-3-bp, and only one BJ displayed a signature of blunt-end ligation, indicating chromothripsis as the underlying mechanism. Although the BJs did not disrupt any disease-causing gene, the clinical features of the patient were compatible with MEF2C haploinsufficiency syndrome. Complex rearrangements were located approximately 2.5-Mb downstream of MEF2C. Therefore, position effects were considered the mechanism of the occurrence of MEF2C haploinsufficiency syndrome.


Assuntos
Transtornos do Neurodesenvolvimento , Translocação Genética , Humanos , Masculino , Lactente , Encéfalo/patologia , Transtornos do Neurodesenvolvimento/genética
3.
Hum Genome Var ; 8(1): 10, 2021 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-33594047

RESUMO

MCT8 deficiency is an X-linked recessive disorder. We report the case of a 2-year-old Japanese boy with MCT8 deficiency caused by a novel frameshift variant, NM_006517.5(SLC16A2_v001):c.966dup [p.(Ile323Hisfs*57)]. He presented no head control and spoke no meaningful words, indicating severe developmental delay. Although missense or in-frame mutations of SLC16A2 are usually related to milder phenotypes and later-onset pyramidal signs, loss-of-function mutations are expected to cause severe clinical symptoms.

4.
Hum Mutat ; 41(12): 2119-2127, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32906213

RESUMO

Chromosomal triplications can be classified into recurrent and nonrecurrent triplications. Most of the nonrecurrent triplications are embedded in duplicated segments, and duplication-inverted triplication-duplication (DUP-TRP/INV-DUP) has been established as one of the mechanisms of triplication. This study aimed to reveal the underlying mechanism of the TRP-DUP-TRP pattern of chromosomal aberrations, in which the appearance of moving averages obtained through array-based comparative genomic hybridization analysis is similar to the shadows of the caldera volcano-like pattern, which were first identified in two patients with neurodevelopmental disabilities. For this purpose, whole-genome sequencing using long-read Nanopore sequencing was carried out to confirm breakpoint junctions. Custom array analysis and Sanger sequencing were also used to detect all breakpoint junctions. As a result, the TRP-DUP-TRP pattern consisted of only two patterns of breakpoint junctions in both patients. In patient 1, microhomologies were identified in breakpoint junctions. In patient 2, more complex architectures with insertional segments were identified. Thus, replication-based mechanisms were considered as a mechanism of the TRP-DUP-TRP pattern.


Assuntos
Quebra Cromossômica , Rearranjo Gênico/genética , Genoma Humano , Pré-Escolar , Aberrações Cromossômicas , Hibridização Genômica Comparativa , Feminino , Dosagem de Genes , Humanos , Lactente , Recém-Nascido , Masculino , Sequenciamento por Nanoporos
5.
Congenit Anom (Kyoto) ; 60(6): 189-193, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32618029

RESUMO

Coffin-Siris syndrome (CSS) is a congenital anomaly syndrome characterized by developmental delay, coarse facial features, and hypoplasia of the fifth digit's nail or phalanges. Herein, we report a case of the 8-year-old female patient who showed developmental delay associated with dysplasia in the macular and large toe area. Comprehensive genomic analysis showed no possible candidate variants, but the subsequent genomic copy number analysis revealed a novel exonic deletion in the coding region of AT-rich interactive domain-containing protein 1B (ARID1B), a gene responsible for CSS. Genomic copy number analysis can aid in diagnosing CSS by confirming undiagnosed exonic deletions in ARID1B. Furthermore, this is the first report of CSS associated with bilateral macular dysplasia.


Assuntos
Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/genética , Proteínas de Ligação a DNA/genética , Éxons , Face/anormalidades , Deformidades Congênitas da Mão/diagnóstico , Deformidades Congênitas da Mão/genética , Deficiência Intelectual/diagnóstico , Deficiência Intelectual/genética , Macula Lutea/anormalidades , Micrognatismo/diagnóstico , Micrognatismo/genética , Pescoço/anormalidades , Fenótipo , Deleção de Sequência , Fatores de Transcrição/genética , Criança , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença , Humanos
6.
Hum Genome Var ; 7: 16, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32509318

RESUMO

Comprehensive genomic analysis was performed in a patient with mild psychomotor developmental delay, elevated creatine kinase, and white matter abnormalities. The results revealed biallelic pathogenic variants in the gene related to merosin-deficient congenital muscular dystrophy, NM_000426.3(LAMA2):c.1338_1339del [p.Gly447Phefs*7] and c.2749 + 2dup, which consist of compound heterozygous involvement with predicted loss-of-function and splicing abnormalities.

7.
Hum Genet ; 139(12): 1555-1563, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32535809

RESUMO

The chromosomal region critical in Down syndrome has long been analyzed through genotype-phenotype correlation studies using data from many patients with partial trisomy 21. Owing to that, a relatively small region of human chromosome 21 (35.9 ~ 38.0 Mb) has been considered as Down syndrome critical region (DSCR). In this study, microarray-based comparative genomic hybridization analysis identified complex rearrangements of chromosome 21 in a patient manifesting clinical features partially overlapped with that of Down syndrome. Although the patient did not show up-slanting palpebral fissures and single transverse palmar creases, other symptoms were consistent with Down syndrome. Rearrangements were analyzed by whole-genome sequencing using Nanopore long-read sequencing. The analysis revealed that chromosome 21 was fragmented into seven segments and reassembled by six connected points. Among 12 breakpoints, 5 are located within the short region and overlapped with repeated segments. The rearrangement resulted in a maximum gain of five copies, but no region showed loss of genomic copy numbers. Breakpoint-junctions showed no homologous region. Based on these findings, chromoanasynthesis was considered as the mechanism. Although the distal 21q22.13 region was not included in the aberrant regions, some of the genes located on the duplicated regions, SOD1, SON, ITSN1, RCAN1, and RUNX1, were considered as possible candidate genes for clinical features of the patient. We discussed the critical region for Down syndrome, with the literature review.


Assuntos
Cromossomos Humanos Par 21/genética , Cromossomos Humanos/genética , Síndrome de Down/genética , Proteínas Adaptadoras de Transporte Vesicular/genética , Aberrações Cromossômicas , Hibridização Genômica Comparativa/métodos , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Proteínas de Ligação a DNA/genética , Síndrome de Down/fisiopatologia , Feminino , Dosagem de Genes/genética , Estudos de Associação Genética , Humanos , Hibridização in Situ Fluorescente/métodos , Lactente , Análise em Microsséries/métodos , Antígenos de Histocompatibilidade Menor/genética , Proteínas Musculares/genética , Superóxido Dismutase-1/genética , Sequenciamento Completo do Genoma
8.
J Hum Genet ; 65(9): 735-741, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32355308

RESUMO

The widespread use of genomic copy number analysis has revealed many previously unknown genomic structural variations, including some which are more complex. In this study, three consecutive microdeletions were identified in the same chromosome by microarray-based comparative genomic hybridization (aCGH) analysis for a patient with a neurodevelopmental disorder. Subsequent fluorescence in situ hybridization (FISH) analyses unexpectedly suggested complicated translocations and inversions. For better understanding of the mechanism, breakpoint junctions were analyzed by nanopore sequencing, as a new long-read whole-genome sequencing (WGS) tool. The results revealed a new chromosomal disruption, giving rise to four junctions in chromosome 7. According the sequencing results of breakpoint junctions, all junctions were considered as the consequence of multiple double-strand breaks and the reassembly of DNA fragments by nonhomologous end-joining, indicating chromothripsis. KMT2E, located within the deletion region, was considered as the gene responsible for the clinical features of the patient. Combinatory usage of aCGH and FISH analyses would be recommended for interpretation of structural variations analyzed through WGS.


Assuntos
Aberrações Cromossômicas , Hibridização Genômica Comparativa/métodos , Proteínas de Ligação a DNA/genética , Genômica/métodos , Hibridização in Situ Fluorescente/métodos , Transtornos do Neurodesenvolvimento/genética , Quebra Cromossômica , Inversão Cromossômica , Quebras de DNA de Cadeia Dupla , Variações do Número de Cópias de DNA , Reparo do DNA por Junção de Extremidades , Feminino , Estudos de Associação Genética , Genoma Humano , Humanos , Sequenciamento por Nanoporos , Análise de Sequência com Séries de Oligonucleotídeos , Inversão de Sequência , Translocação Genética , Sequenciamento Completo do Genoma , Adulto Jovem
9.
Congenit Anom (Kyoto) ; 60(1): 10-14, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30684281

RESUMO

Genomic copy number variations (CNVs) can be detected by chromosomal microarray testing. However, upon final diagnosis, other methods may be recommended for a validation method to confirm CNVs. Trio analyses or carrier detection in family members are also frequently required. Previously, fluorescence in situ hybridization and/or quantitative PCR have been used; however, these methods present limitations. The purpose of this study was to establish a simple and rapid method to detect genomic copy numbers. We utilized droplet digital PCR (dPCR) with an intercalation method. Thirteen patients, who were diagnosed with MECP2 duplications via chromosomal microarray testing, were enrolled in this study. Four of their female relatives, who were verified as carriers of MECP2 duplications, were also included. Genomic copy numbers of MECP2 and IRAK1 were analyzed in comparison with reference genes: XIST and AR on the X-chromosome, and RPP30 and RPPH1 on the autosomal chromosomes. As a result, genomic copy numbers of MECP2 were rapidly and precisely detected by the dPCR system established in this study. This method can be widely applied as a diagnostic method to confirm CNVs on other chromosomal regions.


Assuntos
Duplicação Gênica/genética , Quinases Associadas a Receptores de Interleucina-1/genética , Proteína 2 de Ligação a Metil-CpG/genética , Reação em Cadeia da Polimerase/métodos , Autoantígenos/genética , Aberrações Cromossômicas , Variações do Número de Cópias de DNA/genética , Feminino , Genoma/genética , Humanos , Hibridização in Situ Fluorescente , Masculino , Proteína 2 de Ligação a Metil-CpG/isolamento & purificação , Análise em Microsséries , Linhagem , RNA Longo não Codificante/genética , Receptores Androgênicos/genética , Ribonuclease P/genética
10.
Am J Med Genet A ; 182(3): 521-526, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31821719

RESUMO

Primrose syndrome is a congenital malformation syndrome characterized by intellectual disability, developmental delay, progressive muscle wasting, and ear lobe calcification. Mutations in the ZBTB20 gene have been established as being accountable for this syndrome. In this study, a novel de novo ZBTB20 mutation, NM_001164342.2:c.1945C>T (p.Leu649Phe), has been identified through whole exome sequencing (WES) in a female patient presenting a typical Primrose phenotype. Because the present patient exhibited recurrent otitis media, detailed immunological examinations were performed in this study and subnormal immunoglobulin levels were firstly identified in a Primrose patient. Anatomical anomaly of the inner ear has never been reported in this patient and WES data did not include any relevant variants causally linked with the immunologic defect. Thus, there is a possibility of a relation between an unclassified immunodeficiency with selective IgG2 deficiency and Primrose syndrome and this may be the reason of recurrent otitis media frequently observed in Primrose patients. Because subnormal levels of IgG2 in this patient might be caused by an unrelated and still uncharacterized genetic cause, further studies are required to prove the causal link between aberrant ZBTB20 function and immunodeficiency.


Assuntos
Anormalidades Múltiplas/genética , Calcinose/genética , Deficiências do Desenvolvimento/genética , Otopatias/genética , Predisposição Genética para Doença , Deficiência Intelectual/genética , Atrofia Muscular/genética , Proteínas do Tecido Nervoso/genética , Fatores de Transcrição/genética , Anormalidades Múltiplas/diagnóstico , Anormalidades Múltiplas/patologia , Calcinose/diagnóstico , Calcinose/patologia , Criança , Deficiências do Desenvolvimento/diagnóstico , Deficiências do Desenvolvimento/patologia , Otopatias/diagnóstico , Otopatias/patologia , Feminino , Humanos , Imunoglobulina G/genética , Deficiência Intelectual/diagnóstico , Deficiência Intelectual/patologia , Masculino , Atrofia Muscular/diagnóstico , Atrofia Muscular/patologia , Mutação de Sentido Incorreto/genética , Fenótipo , Sequenciamento do Exoma
11.
Intractable Rare Dis Res ; 8(3): 198-202, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31523598

RESUMO

Pelizaeus-Merzbacher disease (PMD) is an X-linked, recessively inherited disorder associated with hypomyelination in the brain white matter. Mutations involving the proteolipid protein 1 gene (PLP1) located on Xq22.2 are responsible for PMD. PLP1 duplication is the major genetic abnormality in PMD patients. In this study, we utilized droplet-digital polymerase chain reaction (ddPCR) as a potential method to detect PLP1 duplications. Samples from four PMD patients and one of their mothers were used as positive controls. They had been previously diagnosed as having an additional PLP1 copy by chromosomal microarray testing. Genomic copy number of PLP1 was analyzed in triplicate experiments and compared with reference genes XIST and AR on the X-chromosome, and RPP30 and RPPH1 on the autosomes. As a result, precise results were obtained for each triplicate procedure. Thus, we concluded that triplicate experiments are no longer necessary. Compared to other methods, including fluorescence in-situ hybridization, multiplex ligation-dependent probe amplification, chromosomal microarray testing, and quantitative PCR, we were able to establish ddPCR results rapidly with very small amounts of DNA. In conclusion, we showed that ddPCR can be a potential diagnostic tool to confirm genomic copy number as a routine clinical application, including in prenatal diagnostic settings.

12.
Brain Dev ; 41(9): 776-782, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31171384

RESUMO

BACKGROUND: Recently, many genes related to neurodevelopmental disorders have been identified by high-throughput genomic analysis; however, a comprehensive understanding of the mechanism underlying neurodevelopmental disorders remains to be established. To further understand these underlying mechanisms, we performed a comprehensive genomic analysis of patients with undiagnosed neurodevelopmental disorders. METHODS: Genomic analysis using next-generation sequencing with a targeted panel was performed for a total of 133 Japanese patients (male/female, 81/52) with previously undiagnosed neurodevelopmental disorders, including developmental delay (DD), intellectual disability (ID), autism spectrum disorder (ASD), and epilepsy. Genomic copy numbers were also analyzed using the eXome Hidden Markov Model (XHMM). RESULTS: Thirty-nine patients (29.3%) exhibited pathogenic or likely pathogenic findings with single-gene variants or chromosomal aberrations. Among them, 20 patients were presented here. Pathogenic or likely pathogenic variants were identified in 18 genes, including ACTG1, CACNA1A, CHD2, CDKL5, DNMT3A, EHMT1, GABRB3, GABRG2, GRIN2B, KCNQ3, KDM5C, MED13L, SCN2A, SHANK3, SMARCA2, STXBP1, SYNGAP1, and TBL1XR1. CONCLUSION: A diagnostic yield of 29.3% in this study was nearly the same as that previously reported from other countries. Thus, we suggest that there is no difference in genomic backgrounds in Japanese patients with undiagnosed neurodevelopmental disabilities. Although most of the patients possessed de novo variants, one of the patients showed an X-linked inheritance pattern. As X-linked recessive disorders exhibit the possibility of recurrent occurrence in the family, comprehensive molecular diagnosis is important for genetic counseling.


Assuntos
Transtornos do Neurodesenvolvimento/genética , Povo Asiático/genética , Pré-Escolar , Variações do Número de Cópias de DNA , Feminino , Predisposição Genética para Doença , Humanos , Japão , Masculino , Polimorfismo de Nucleotídeo Único
13.
J Hum Genet ; 64(7): 665-671, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31004103

RESUMO

Pelizaeus-Merzbacher disease (PMD) is an X-linked recessive disorder caused by abnormalities in the gene PLP1. Most females harboring heterozygous PLP1 abnormalities are basically asymptomatic. However, as a result of abnormal patterns of X-chromosome inactivation, it is possible for some female carriers to be symptomatic. Whole-exome sequencing of a female patient with unknown spastic paraplegia was performed to obtain a molecular diagnosis. As a result, a de novo heterozygous single-nucleotide deletion in PLP1 [NM_000533.5(PLP1_v001):c.783del; p.Thr262Leufs*20] was identified. RNA sequencing was performed in a patient-derived lymphoblastoid cell line, confirming mono-allelic expression of the mutated allele and abnormal inactivation of the wild-type allele. The patient-derived lymphoblastoid cell line was then treated with VX680 or 5azadC, which resulted in restored expression of the wild-type allele. These two agents thus have the potential to reverse inappropriately-skewed inactivation of the X-chromosome.


Assuntos
Mutação da Fase de Leitura , Proteína Proteolipídica de Mielina/genética , Paraplegia/genética , Doença de Pelizaeus-Merzbacher/genética , Alelos , Linhagem Celular , Criança , Cromossomos Humanos X/genética , Cromossomos Humanos X/metabolismo , Decitabina/farmacologia , Feminino , Humanos , Doença de Pelizaeus-Merzbacher/patologia , Doença de Pelizaeus-Merzbacher/terapia , Piperazinas/farmacologia , Sequenciamento do Exoma
15.
Intractable Rare Dis Res ; 7(4): 245-250, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30560016

RESUMO

Because biallelic SZT2 variants have been reported in patients with neurodevelopmental disorders associated with various degrees of developmental delay, intractable seizures, and distinctive features; this condition is recognized as an autosomal recessive disorder. Previously, eleven patients have been reported and most of them have compound heterozygous SZT2 variants, leading to premature termination. In these patients, all reported variants were unique and there were no common pathogenic variants identified. In this study, we identified a paternal uniparental disomy of chromosome 1 in a patient with a neurodevelopmental disorder associated with severe intellectual disability, intractable epilepsy, autistic features, distinctive features, and transient macrocephaly. This resulted in homozygous patterns through chromosome 1. Among the variants in chromosome 1, a rare SZT2 variant, NM_015284.3:c.6553C>T (p.Arg2185Trp), was selected as a powerful candidate variant in this patient. Although the clinical features of this patient are relatively milder than that reported previously, it may be derived from genetic heterogeneity. This is the first report of a homozygous missense SZT2 variant.

16.
Am J Med Genet A ; 176(12): 2803-2807, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30152144

RESUMO

Vici syndrome is a rare, autosomal recessive, multisystem disorder, characterized by agenesis of the corpus callosum, cataracts, psychomotor delay, cardiomyopathy, hypopigmentation, and recurrent infections. Mutations in the ectopic P-granules autophagy protein 5 homolog gene (EPG5), which encodes a key autophagy regulator, are responsible for this syndrome. A 3-year-old Japanese girl manifesting similar symptoms to those found in patients with Vici syndrome showed intractable diarrhea, rather than immunodeficiency. Whole exome sequencing identified only a heterozygous variant in EPG5, NM_020964.2(EPG5):c.3389A > C (p.His1130Pro), which was inherited from her mother. Sequencing analyses of the EPG5 messenger RNA showed only an altered nucleotide "C" at position, c.3389, indicating decreased expression of the wild-type allele. Microarray-based comparative genomic hybridization revealed a de novo microduplication in the exon 1 region. Large exon deletions and duplications of EPG5 have never been reported so far. This was considered the cause of the decreased expression of the wild-type allele. In conclusion, we successfully identified novel compound heterozygous mutations in EPG5 in a patient who was clinically considered to have Vici syndrome.


Assuntos
Agenesia do Corpo Caloso/diagnóstico , Agenesia do Corpo Caloso/genética , Catarata/diagnóstico , Catarata/genética , Éxons , Duplicação Gênica , Heterozigoto , Proteínas de Membrana Lisossomal/genética , Mutação , Proteínas de Transporte Vesicular/genética , Proteínas Relacionadas à Autofagia , Encéfalo/anormalidades , Encéfalo/diagnóstico por imagem , Pré-Escolar , Feminino , Estudos de Associação Genética , Testes Genéticos , Genômica/métodos , Humanos , Japão , Imageamento por Ressonância Magnética , Fenótipo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...