Browsing by Subject "Mosaicism"

Now showing 1 - 5 of 5
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    Analysis of integration sites of transgenic sheep generated by lentiviral vectors using next-generation sequencing technology
    (2014-07-31) Chen, Yu-Hsiang; Malkova, Anna; Cornetta, Kenneth; Randall, Stephen Karl, 1953-; Atkinson, Simon
    The development of new methods to carry out gene transfer has many benefits to several fields, such as gene therapy, agriculture and animal health. The newly established lentiviral vector systems further increase the efficiency of gene transfer dramatically. Some studies have shown that lentiviral vector systems enhance efficiency over 10-fold higher than traditional pronuclear injection. However, the timing for lentiviral vector integration to occur remains unclear. Integrating in different stages of embryogenesis might lead to different integration patterns between tissues. Moreover, in our previous study we found that the vector copy number in transgenic sheep varied, some having one or more copies per cells while other animals having less than one copy per cell suggesting mosaicism. Here I hypothesized that injection of a lentiviral vector into a single cell embryo can lead to integration very early in embryogenesis but can also occur after several cell divisions. In this study, we focus on investigating integration sites in tissues developing from different germ layers as well as extraembryonic tissues to determine when integration occurs. In addition, we are also interested in insertional mutagenesis caused by viral sequence integration in or near gene regions. We utilize linear amplification-mediated polymerase chain reaction (LAM-PCR) and next- generation sequencing (NGS) technology to determine possible integration sites. In this study, we found the evidence based on a series of experiments to support my hypothesis, suggesting that integration event also happens after several cell divisions. For insertional mutagenesis analysis, the closest genes can be found according to integration sites, but they are likely too far away from the integration sites to be influenced. A well-annotated sheep genome database is needed for insertional mutagenesis analysis.
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    Cytogenetic studies of mosaicism
    (1987) Higgins, Michael D.
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    Genetic mosaicism, intrafamilial phenotypic heterogeneity, and molecular defects of a novel missense SLC6A1 mutation associated with epilepsy and ADHD
    (Elsevier, 2021) Poliquin, Sarah; Hughes, Inna; Shen, Wangzhen; Mermer, Felicia; Wang, Juexin; Mack, Taralynn; Xu, Dong; Kang, Jing-Qiong; Medical and Molecular Genetics, School of Medicine
    Background: Mutations in SLC6A1, encoding γ-aminobutyric acid (GABA) transporter 1 (GAT-1), have been recently associated with a spectrum of neurodevelopmental disorders ranging from variable epilepsy syndromes, intellectual disability (ID), autism and others. To date, most identified mutations are de novo. We here report a pedigree of two siblings associated with myoclonic astatic epilepsy, attention deficit hyperactivity disorder (ADHD), and ID. Methods: Next-generation sequencing identified a missense mutation in the SLC6A1 gene (c.373G > A(p.Val125Met)) in the sisters but not in their shared mother who is also asymptomatic, suggesting gonadal mosaicism. We have thoroughly characterized the clinical phenotypes: EEG recordings identified features for absence seizures and prominent bursts of occipital intermittent rhythmic delta activity (OIRDA). The molecular pathophysiology underlying the clinical phenotypes was assessed using a multidisciplinary approach including machine learning, confocal microscopy, and high-throughput 3H radio-labeled GABA uptake assays in mouse astrocytes and neurons. Results: The GAT-1(Val125Met) mutation destabilizes the global protein conformation and reduces transporter protein expression at total and cell surface. The mutant transporter protein was localized intracellularly inside the endoplasmic reticulum (ER) in both HEK293T cells and astrocytes which may directly contribute to seizures in patients. Radioactive 3H-labeled GABA uptake assay indicated the mutation reduced the function of the mutant GAT-1(Val125Met) to ~30% of the wildtype. Conclusions: The seizure phenotypes, ADHD, and impaired cognition are likely caused by a partial loss-of-function of GAT-1 due to protein destabilization resulting from the mutation. Reduced GAT-1 function in astrocytes and neurons may consequently alter brain network activities such as increased seizures and reduced attention.
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    Hormonal suppression of mini-puberty in a neonate with mosaic 45X/46XY disorder of sexual development
    (Elsevier, 2020-05-03) Kaefer, Martin; Eugster, Erica; Medicine, School of Medicine
    Disorders of Sex Development (DSD) are some of the most controversial and challenging conditions that pediatric urologists treat. This may be especially true in mosaic 45X/46XY DSD, due to the inability to ascertain in the neonatal period which gender identity will best suit a given child with this condition. It has therefore been proposed to forgo any irreversible surgical interventions. In order to address the concern of early testosterone production in a nonsurgical manner we describe a case in which we treat a patient with a GnRH agonist to block the early physiologic rise in testosterone during the neonatal mini-puberty.
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    Maternal mosaicism in long QT syndrome due to a pathogenic variant in KCNH2
    (Elsevier, 2020-11-16) Sawyer, Briana L.; Tristani-Firouzi, Martin; Wells, Layne E.; Vatta, Matteo; Etheridge, Susan P.; Medical and Molecular Genetics, School of Medicine