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The Indian Journal of Medical Research Sep 2019Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder and is caused mainly by deletion, duplication and point mutations in the DMD gene. Diagnosis of DMD...
BACKGROUND & OBJECTIVES
Duchenne muscular dystrophy (DMD) is an X-linked recessive disorder and is caused mainly by deletion, duplication and point mutations in the DMD gene. Diagnosis of DMD has been a challenge as the mutations in the.
DMD
gene are heterogeneous and require more than one diagnostic strategy for the validation of the mutation. This study was planned to evaluate the targeted next-generation sequencing (NGS) as a single platform to detect all types of mutations in the DMD gene, thereby reducing the time and costs compared to conventional sequential testing and also provide precise genetic information for emerging gene therapies.
METHODS
The study included 20 unrelated families and 22 patients from an Indian population who were screened for DMD based on phenotypes such as scoliosis, toe walking and loss of ambulation. Peripheral blood DNA was isolated and subjected to multiplex ligation-dependent probe amplification (MLPA) and targeted NGS of the DMD gene to identify the nature of the mutation.
RESULTS
In the study patients, 77 per cent of large deletion mutations and 23 per cent single-nucleotide variations (SNVs) were identified. Novel mutations were also identified along with reported deletions, point mutations and partial deletions within the exon of the DMD gene.
INTERPRETATION & CONCLUSIONS
Our findings showed the importance of NGS in the routine diagnostic practice in the identification of DMD mutations over sequential testing. It may be used as a single-point diagnostic strategy irrespective of the mutation type, thereby reducing the turnaround time and cost for multiple diagnostic tests such as MLPA and Sanger sequencing. Though MLPA is a sensitive technique and is the first line of a diagnostic test, the targeted NGS of the DMD gene may have an advantage of having a single diagnostic test. A study on a larger number of patients is needed to highlight NGS as a single, comprehensive platform for the diagnosis of DMD.
Topics: Adolescent; Child; Child, Preschool; DNA Mutational Analysis; Dystrophin; Exons; Gene Deletion; Genome, Human; High-Throughput Nucleotide Sequencing; Humans; India; Male; Multiplex Polymerase Chain Reaction; Muscular Dystrophy, Duchenne; Mutation; Phenotype; Point Mutation; Polymorphism, Single Nucleotide; Sequence Deletion
PubMed: 31719299
DOI: 10.4103/ijmr.IJMR_290_18 -
Genes Jan 2020Understanding mitochondrial DNA (mtDNA) evolution and inheritance has broad implications for animal speciation and human disease models. However, few natural models...
Understanding mitochondrial DNA (mtDNA) evolution and inheritance has broad implications for animal speciation and human disease models. However, few natural models exist that can simultaneously represent mtDNA transmission bias, mutation, and copy number variation. Certain isolates of the nematode harbor large, naturally-occurring mtDNA deletions of several hundred basepairs affecting the () gene that can be functionally detrimental. These deletion variants can behave as selfish DNA elements under genetic drift conditions, but whether all of these large deletion variants are transmitted in the same preferential manner remains unclear. In addition, the degree to which transgenerational mtDNA evolution profiles are shared between isolates that differ in their propensity to accumulate the deletion is also unclear. We address these knowledge gaps by experimentally bottlenecking two isolates of with different deletion frequencies for up to 50 generations and performing total DNA sequencing to identify mtDNA variation. We observed multiple mutation profile differences and similarities between isolates, a potentially species-specific pattern of copy number dysregulation, and some evidence for genetic hitchhiking in the deletion-bearing isolate. Our results further support as a practical model for characterizing naturally-occurring mtgenome variation and contribute to the understanding of how mtgenome variation persists in animal populations and how it presents in mitochondrial disease states.
Topics: Animals; Bacterial Proteins; Base Sequence; Caenorhabditis; DNA Copy Number Variations; DNA, Mitochondrial; Gene Deletion; Genetic Variation; Genome, Mitochondrial; Mitochondria; Mutation; NADH Dehydrogenase; Phylogeny; Repetitive Sequences, Nucleic Acid; Sequence Analysis, DNA; Sequence Deletion
PubMed: 31936803
DOI: 10.3390/genes11010077 -
Genetics May 2003The rate of mutation for nucleotide substitution is generally higher among males than among females, likely owing to the larger number of DNA replications in...
The rate of mutation for nucleotide substitution is generally higher among males than among females, likely owing to the larger number of DNA replications in spermatogenesis than in oogenesis. For insertion and deletion (indel) mutations, data from a few human genetic disease loci indicate that the two sexes may mutate at similar rates, possibly because such mutations arise in connection with meiotic crossing over. To address origin- and sex-specific rates of indel mutation we have conducted the first large-scale molecular evolutionary analysis of indels in noncoding DNA sequences from sex chromosomes. The rates are similar on the X and Y chromosomes of primates but about twice as high on the avian Z chromosome as on the W chromosome. The fact that indels are not uncommon on the nonrecombining Y and W chromosomes excludes meiotic crossing over as the main cause of indel mutation. On the other hand, the similar rates on X and Y indicate that the number of DNA replications (higher for Y than for X) is also not the main factor. Our observations are therefore consistent with a role of both DNA replication and recombination in the generation of short insertion and deletion mutations. A significant excess of deletion compared to insertion events is observed on the avian W chromosome, consistent with gradual DNA loss on a nonrecombining chromosome.
Topics: Animals; Birds; Female; Humans; Introns; Male; Molecular Sequence Data; Point Mutation; Primates; Sequence Analysis, DNA; Sequence Deletion; Sex Chromosomes
PubMed: 12750337
DOI: 10.1093/genetics/164.1.259 -
Acta Biochimica Polonica Aug 2021We present here an alternative for two-promoter systems ensuring highly diverse expression of several genes from a single promoter. This approach assumes an introduction...
We present here an alternative for two-promoter systems ensuring highly diverse expression of several genes from a single promoter. This approach assumes an introduction of a deletion mutation into an A/T homopolymeric run in a gene's proximal part, and employs the transcriptional slippage mechanism for insertion-dependent reinstatement of the proper reading frame by the T7 RNA polymerase.
Topics: Base Sequence; DNA-Directed RNA Polymerases; Escherichia coli; Gene Expression; Genetic Techniques; Promoter Regions, Genetic; RNA, Messenger; Sequence Deletion; Transcription, Genetic; Viral Proteins
PubMed: 34436838
DOI: 10.18388/abp.2020_5736 -
BMC Medical Genetics Jan 2019Autosomal recessive nail dysplasia is characterized by thick and hard nails with a very slow growth on the hands and feet. Mutations in FZD6 gene were found to be...
BACKGROUND
Autosomal recessive nail dysplasia is characterized by thick and hard nails with a very slow growth on the hands and feet. Mutations in FZD6 gene were found to be associated with autosomal recessive nail dysplasia in 2011. Presently, only seven mutations have been reported in FZD6 gene; five mutations are clustered in the C-terminus, one is at the seventh transmembrane domain, and another is at the very beginning of third extracellular loop.
METHODS
Whole exome sequencing (WES) was applied to the index case, her one affected sister and her healthy consanguineous parents. The mutation was verified via Sanger sequencing. Molecular dynamics simulations of the predicted structures of native and mutant proteins were compared to gain insight into the pathogenicity mechanism of the mutation.
RESULTS
Here, we report a homozygous 8 bp deletion mutation, p.Gly559Aspfs*16; c.1676_1683delGAACCAGC, in FZD6 gene which causes a frameshift and creates a premature stop codon at position 16 of the new reading frame. Our molecular dynamics calculations predict that the pathogenicity of this frameshift mutation may be caused by the change in entropy of the protein with negative manner, disturbing the C-terminal domain structure, and hence interaction partners of FZD6.
CONCLUSION
We identified a homozygous deletion mutation in FZD6 in a consanguineous Turkish family with nail dysplasia. We also provide a molecular mechanism about the effects of the deletion on the protein structure and its possible motions. This study provides a pathogenicity mechanism for this mutation in nail dysplasia for the first time.
Topics: Adult; Amino Acid Sequence; Codon, Nonsense; Consanguinity; Female; Frameshift Mutation; Frizzled Receptors; Genetic Association Studies; Genetic Predisposition to Disease; Homozygote; Humans; Models, Molecular; Mutation; Nails, Malformed; Pedigree; Protein Conformation; Sequence Analysis; Sequence Deletion; Turkey
PubMed: 30642273
DOI: 10.1186/s12881-019-0746-6 -
Electrophoresis Oct 2018Baxα, a key tumor suppressor gene, will not be expressed correctly as a result of single nucleotide mutation in its microsatellite region; Instead, BaxΔ2, an isoform...
Baxα, a key tumor suppressor gene, will not be expressed correctly as a result of single nucleotide mutation in its microsatellite region; Instead, BaxΔ2, an isoform of Baxα, is often produced. In addition, lack of the exon 2 due to an alternative splicing, BaxΔ2 has the same sequence as Baxα except single base deletion from eight continuous guanines (G8) to G7. Most of the currently available methods for Bax∆2 detection are inefficient and time-consuming, and/or require the use of labels or dyes. In this work, we reported a label-free nanopore sensing strategy to differentiate between Baxα and BaxΔ2 with a DNA polymer as a molecular probe based on alternative spliced sequences. Two DNA molecules were designed to selectively detect Baxα and BaxΔ2, respectively. The method was rapid, accurate, and highly sensitive: picomolar concentrations of target nucleic acids could be detected in minutes. Our developed simple and fast nanopore-based detection strategy is not only useful for distinguishing between Baxα and Bax∆2, but also provides a useful tool for detection of other single-base mutations in genetic diagnosis.
Topics: DNA; DNA Mutational Analysis; Humans; Limit of Detection; Molecular Diagnostic Techniques; Molecular Probe Techniques; Nanopores; Sequence Deletion; bcl-2-Associated X Protein
PubMed: 29998460
DOI: 10.1002/elps.201800193 -
European Journal of Biochemistry Dec 1999Mutations in the LIS1 gene may result in severe abnormalities of brain cortical layering known as lissencephaly. Most lissencephaly-causing LIS1 mutations are deletions...
Mutations in the LIS1 gene may result in severe abnormalities of brain cortical layering known as lissencephaly. Most lissencephaly-causing LIS1 mutations are deletions that encompass the entire gene, therefore the mechanism of the disease is regarded as haploinsufficiency. So far, 13 different intragenic mutations have been reported: one point mutation, H149R; deletion of exon 9, which results in deleted acids Delta301-334; deletion of exon 4, which results in deleted amino acids Delta40-64; 10 mutations resulting in truncated proteins and one predicted to result in extra amino acids. We studied the consequences of the point mutation, deletion mutation and one of the reported truncations. In order to study LIS1 structure function, we introduced an additional point mutation and other truncations in different regions of the protein. The consequences of these mutations to protein folding were studied by gel filtration, sucrose density gradient centrifugation and measuring resistance to trypsin cleavage. On the basis of our results, we suggest that all truncation mutations and lissencephaly-causing point mutations or internal deletion result in a reduction in the amount of correctly folded LIS1 protein.
Topics: 1-Alkyl-2-acetylglycerophosphocholine Esterase; Amino Acid Sequence; Animals; Brain; Cattle; Computer Simulation; Female; Humans; Male; Mice; Microtubule-Associated Proteins; Models, Molecular; Molecular Sequence Data; Mutagenesis, Site-Directed; Mutation; Point Mutation; Protein Folding; Recombinant Proteins; Sequence Deletion; Sequence Homology, Amino Acid
PubMed: 10583396
DOI: 10.1046/j.1432-1327.1999.00942.x -
Human Mutation Mar 2022Microdeletions and gross deletions are important causes (~20%) of human inherited disease and their genomic locations are strongly influenced by the local DNA sequence...
Microdeletions and gross deletions are important causes (~20%) of human inherited disease and their genomic locations are strongly influenced by the local DNA sequence environment. This notwithstanding, no study has systematically examined their underlying generative mechanisms. Here, we obtained 42,098 pathogenic microdeletions and gross deletions from the Human Gene Mutation Database (HGMD) that together form a continuum of germline deletions ranging in size from 1 to 28,394,429 bp. We analyzed the DNA sequence within 1 kb of the breakpoint junctions and found that the frequencies of non-B DNA-forming repeats, GC-content, and the presence of seven of 78 specific sequence motifs in the vicinity of pathogenic deletions correlated with deletion length for deletions of length ≤30 bp. Further, we found that the presence of DR, GQ, and STR repeats is important for the formation of longer deletions (>30 bp) but not for the formation of shorter deletions (≤30 bp) while significantly (χ , p < 2E-16) more microhomologies were identified flanking short deletions than long deletions (length >30 bp). We provide evidence to support a functional distinction between microdeletions and gross deletions. Finally, we propose that a deletion length cut-off of 25-30 bp may serve as an objective means to functionally distinguish microdeletions from gross deletions.
Topics: Base Composition; Base Sequence; DNA; Genome, Human; Humans; Mutation; Sequence Deletion
PubMed: 34918412
DOI: 10.1002/humu.24314 -
Nucleic Acids Research Oct 2016The accumulation of somatic mitochondrial DNA (mtDNA) mutations contributes to the pathogenesis of human disease. Currently, mitochondrial mutations are largely...
The accumulation of somatic mitochondrial DNA (mtDNA) mutations contributes to the pathogenesis of human disease. Currently, mitochondrial mutations are largely considered results of inaccurate processing of its heavily damaged genome. However, mainly from a lack of methods to monitor mtDNA mutations with sufficient sensitivity and accuracy, a link between mtDNA damage and mutation has not been established. To test the hypothesis that mtDNA-damaging agents induce mtDNA mutations, we exposed MutaMouse mice to benzo[a]pyrene (B[a]P) or N-ethyl-N-nitrosourea (ENU), daily for 28 consecutive days, and quantified mtDNA point and deletion mutations in bone marrow and liver using our newly developed Digital Random Mutation Capture (dRMC) and Digital Deletion Detection (3D) assays. Surprisingly, our results demonstrate mutagen treatment did not increase mitochondrial point or deletion mutation frequencies, despite evidence both compounds increase nuclear DNA mutations and demonstrated B[a]P adduct formation in mtDNA. These findings contradict models of mtDNA mutagenesis that assert the elevated rate of mtDNA mutation stems from damage sensitivity and abridged repair capacity. Rather, our results demonstrate induced mtDNA damage does not readily convert into mutation. These findings suggest robust mitochondrial damage responses repress induced mutations after mutagen exposure.
Topics: Animals; Benzo(a)pyrene; Bone Marrow; Cell Nucleus; DNA Adducts; DNA, Mitochondrial; Ethylnitrosourea; Liver; Male; Mice; Mutagenesis; Mutagens; Point Mutation; Sequence Deletion
PubMed: 27550180
DOI: 10.1093/nar/gkw716 -
Breast Cancer Research : BCR 2003Germline mutations in the human checkpoint gene, hCHK2, were first identified in 1999 in cases of Li-Fraumeni syndrome. Recent studies have demonstrated that the hCHK2... (Review)
Review
Germline mutations in the human checkpoint gene, hCHK2, were first identified in 1999 in cases of Li-Fraumeni syndrome. Recent studies have demonstrated that the hCHK2 1100delC mutation acts as a low-penetrance tumour suppressor gene in familial breast cancer not associated with mutations in BRCA1 or BRCA2. The present article describes the published studies on hCHK2 1100delC and addresses some of the key questions raised.
Topics: Base Sequence; Breast Neoplasms; Checkpoint Kinase 2; Cytosine; Genes, Tumor Suppressor; Humans; Protein Kinases; Protein Serine-Threonine Kinases; Risk Factors; Sequence Deletion
PubMed: 12793891
DOI: 10.1186/bcr582