scholarly journals Functional reassessment of PAX6 single nucleotide variants by in vitro splicing assay

2018 ◽  
Vol 27 (3) ◽  
pp. 488-493 ◽  
Author(s):  
Alexandra Yu. Filatova ◽  
Tatiana A. Vasilyeva ◽  
Andrey V. Marakhonov ◽  
Anna A. Voskresenskaya ◽  
Rena A. Zinchenko ◽  
...  
Keyword(s):  
Single Nucleotide Variants ◽  
Single Nucleotide ◽  
In Vitro Splicing

scholarly journals Association between HOX Transcript Antisense RNA Single-Nucleotide Variants and Recurrent Implantation Failure

2021 ◽  
Vol 22 (6) ◽  
pp. 3021
Author(s):  
Jeong Yong Lee ◽  
Eun Hee Ahn ◽  
Hyeon Woo Park ◽  
Ji Hyang Kim ◽  
Young Ran Kim ◽  
...  
Keyword(s):  
Antisense Rna ◽  
Coagulation Factors ◽  
Healthy Controls ◽  
Implantation Failure ◽  
Single Nucleotide Variants ◽  
Recurrent Implantation Failure ◽  
Single Nucleotide ◽  
Vitro Fertilization ◽  
And Control

Recurrent implantation failure (RIF) refers to the occurrence of more than two failed in vitro fertilization–embryo transfers (IVF-ETs) in the same individual. RIF can occur for many reasons, including embryo characteristics, immunological factors, and coagulation factors. Genetics can also contribute to RIF, with some single-nucleotide variants (SNVs) reported to be associated with RIF occurrence. We examined SNVs in a long non-coding RNA, homeobox (HOX) transcript antisense RNA (HOTAIR), which is known to affect cancer development. HOTAIR regulates epigenetic outcomes through histone modifications and chromatin remodeling. We recruited 155 female RIF patients and 330 healthy controls, and genotyped HOTAIR SNVs, including rs4759314, rs920778, rs7958904, and rs1899663, in all participants. Differences in these SNVs were compared between the patient and control groups. We identified significant differences in the occurrence of heterozygous genotypes and the dominant expression model for the rs1899663 and rs7958904 SNVs between RIF patients and control subjects. These HOTAIR variants were associated with serum hemoglobin (Hgb), luteinizing hormone (LH), total cholesterol (T. chol), and blood urea nitrogen (BUN) levels, as assessed by analysis of variance (ANOVA). We analyzed the four HOTAIR SNVs and found significant differences in haplotype patterns between RIF patients and healthy controls. The results of this study showed that HOTAIR is not only associated with the development of cancer but also with pregnancy-associated diseases. This study represents the first report showing that HOTAIR is correlated with RIF.

scholarly journals An efficient and adaptable workflow for editing disease-relevant single nucleotide variants using CRISPR/Cas9

2021 ◽  
Author(s):  
Inga Usher ◽  
Lorena Ligammari ◽  
Sara Ahrabi ◽  
Emily Hepburn ◽  
Calum Connolly ◽  
...  
Keyword(s):  
Genetic Research ◽  
Bone Cancer ◽  
Illumina Miseq ◽  
Genetic Alterations ◽  
Single Nucleotide Variants ◽  
Experimental Conditions ◽  
Single Nucleotide ◽  
Droplet Pcr ◽  
Low Efficiency

Single nucleotide variants are the commonest genetic alterations in the human genome. At least 60,000 have been reported to be associated with disease. The CRISPR/Cas9 system has transformed genetic research, making it possible to edit single nucleotides and study the function of genetic variants in vitro. While significant advances have improved the efficiency of CRISPR/Cas9, the editing of single nucleotides remains challenging. There are two major obstacles: low efficiency of accurate editing and the isolation of these cells from a pool of cells with other editing outcomes. We present data from 85 transfections of induced pluripotent stem cells and an immortalised cell line, comparing the effects of altering CRISPR/Cas9 design and experimental conditions on rates of single nucleotide substitution. We targeted variants in TP53, which predispose to several cancers, and in TBXT which is implicated in the pathogenesis of the bone cancer, chordoma. We describe a scalable and adaptable workflow for single nucleotide editing that incorporates contemporary techniques including Illumina MiSeq sequencing, TaqMan qPCR and digital droplet PCR for screening transfected cells as well as quality control steps to mitigate against common pitfalls. This workflow can be applied to CRISPR/Cas9 and other genome editing systems to maximise experimental efficiency.

scholarly journals Rare single-nucleotide DAB1 variants and their contribution to Schizophrenia and autism spectrum disorder susceptibility

2020 ◽  
Vol 7 (1) ◽  
Author(s):  
Yoshihiro Nawa ◽  
Hiroki Kimura ◽  
Daisuke Mori ◽  
Hidekazu Kato ◽  
Miho Toyama ◽  
...  
Keyword(s):  
Rare Variants ◽  
Adaptor Protein ◽  
Autism Spectrum ◽  
Case Group ◽  
Missense Mutations ◽  
Single Nucleotide Variants ◽  
Single Nucleotide ◽  
Next Generation Sequencing Technology ◽  
A Minor

AbstractDisabled 1 (DAB1) is an intracellular adaptor protein in the Reelin signaling pathway and plays an essential role in correct neuronal migration and layer formation in the developing brain. DAB1 has been repeatedly reported to be associated with neurodevelopmental disorders including schizophrenia (SCZ) and autism spectrum disorders (ASD) in genetic, animal, and postmortem studies. Recently, increasing attention has been given to rare single-nucleotide variants (SNVs) found by deep sequencing of candidate genes. In this study, we performed exon-targeted resequencing of DAB1 in 370 SCZ and 192 ASD patients using next-generation sequencing technology to identify rare SNVs with a minor allele frequency <1%. We detected two rare missense mutations (G382C, V129I) and then performed a genetic association study in a sample comprising 1763 SCZ, 380 ASD, and 2190 healthy control subjects. Although no statistically significant association with the detected mutations was observed for either SCZ or ASD, G382C was found only in the case group, and in silico analyses and in vitro functional assays suggested that G382C alters the function of the DAB1 protein. The rare variants of DAB1 found in the present study should be studied further to elucidate their potential functional relevance to the pathophysiology of SCZ and ASD.

scholarly journals Multi-omics analysis to identify driving factors in colorectal cancer

Epigenomics ◽  
2020 ◽  
Vol 12 (18) ◽  
pp. 1633-1650
Author(s):  
Xi Xu ◽  
Chaoju Gong ◽  
Yunfeng Wang ◽  
Yanyan Hu ◽  
Hong Liu ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Molecular Mechanisms ◽  
Copy Number Variations ◽  
The Cancer Genome Atlas ◽  
Single Nucleotide Variants ◽  
Single Nucleotide ◽  
Omics Analysis ◽  
Competing Endogenous Rna Network ◽  
Cancer Genome Atlas

Aim: We aim to identify driving genes of colorectal cancer (CRC) through multi-omics analysis. Materials & methods: We downloaded multi-omics data of CRC from The Cancer Genome Atlas dataset. Integrative analysis of single-nucleotide variants, copy number variations, DNA methylation and differentially expressed genes identified candidate genes that carry CRC risk. Kernal genes were extracted from the weighted gene co-expression network analysis. A competing endogenous RNA network composed of CRC-related genes was constructed. Biological roles of genes were further investigated in vitro. Results: We identified LRRC26 and REP15 as novel prognosis-related driving genes for CRC. LRRC26 hindered tumorigenesis of CRC in vitro. Conclusion: Our study identified novel driving genes and may provide new insights into the molecular mechanisms of CRC.

scholarly journals Effects of FSHR and FSHB Variants on Hormonal Profile and Reproductive Outcomes of Infertile Women With Endometriosis

2021 ◽  
Vol 12 ◽  
Author(s):  
Bianca Bianco ◽  
Flavia Altheman Loureiro ◽  
Camila Martins Trevisan ◽  
Carla Peluso ◽  
Denise Maria Christofolini ◽  
...  
Keyword(s):  
Cross Sectional Study ◽  
Reproductive Outcomes ◽  
Single Nucleotide Variants ◽  
Cross Sectional ◽  
Single Nucleotide ◽  
Hormonal Profile ◽  
Ivf Outcomes ◽  
Mild Disease ◽  
Dimensionality Reduction Method

BackgroundSingle nucleotide variants (SNVs) FSHB:c.-211G&gt;T, FSHR:c.919G&gt;A, and FSHR:c.2039G&gt;A were reported to be associated with the variability in FSH and LH levels, and in vitro fertilization (IVF) outcomes. In this study, we aimed to evaluate the effects of FSHB:c.-211G&gt;T, FSHR:c.919G&gt;A, and FSHR:c.2039G&gt;A variants, alone and combined, on the hormonal profile and reproduction outcomes of women with endometriosis.MethodsA cross-sectional study was performed comprising 213 infertile Brazilian women with endometriosis who underwent IVF treatment. Genotyping was performed using TaqMan real-time PCR. Variables were compared according to the genotypes of each variant and genetic models, and the combined effects of the SNVs were evaluated using the multifactorial dimensionality reduction method.ResultsFSHB:c.-211G&gt;T affected LH levels in women with overall endometriosis and minimal/mild disease. FSHR:c.919G&gt;A affected FSH levels in women with overall endometriosis and the number of oocytes retrieved in those with moderate/severe endometriosis. Moreover, the FSHR:c.2039G&gt;A affected FSH levels in women with overall endometriosis, LH levels and total amount of rFSH in those with minimal/mild disease, and number of follicles and number of oocytes retrieved in those with moderate/severe endometriosis. No effect on hormone profile or reproductive outcomes was observed when the genotypes were combined.ConclusionsVariants of the FSHB and FSHR genes separately interfered with the hormonal profiles and IVF outcomes of women with endometriosis.

scholarly journals In vitro zinc supplementation alters synaptic deficits caused by autism spectrum disorder-associated Shank2 point mutations in hippocampal neurons

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Yukti Vyas ◽  
Yewon Jung ◽  
Kevin Lee ◽  
Craig C. Garner ◽  
Johanna M. Montgomery
Keyword(s):  
Hippocampal Neurons ◽  
Fluorescent Protein ◽  
Zinc Supplementation ◽  
Point Mutations ◽  
Autism Spectrum ◽  
Single Nucleotide Variants ◽  
Single Nucleotide ◽  
Green Fluorescent ◽  
The Impact

AbstractAutism Spectrum Disorders (ASDs) are neurodevelopmental disorders characterised by deficits in social interactions and repetitive behaviours. ASDs have a strong genetic basis with mutations involved in the development and function of neural circuitry. Shank proteins act as master regulators of excitatory glutamatergic synapses, and Shank mutations have been identified in people with ASD. Here, we have investigated the impact of ASD-associated Shank2 single nucleotide variants (SNVs) at the synaptic level, and the potential of in vitro zinc supplementation to prevent synaptic deficits. Dissociated rat hippocampal cultures expressing enhanced green fluorescent protein (EGFP) tagged Shank2-Wildtype (WT), and ASD-associated Shank2 single nucleotide variants (SNVs: S557N, V717F, and L1722P), were cultured in the absence or presence of 10 μM zinc. In comparison to Shank2-WT, ASD-associated Shank2 SNVs induced significant decreases in synaptic density and reduced the frequency of miniature excitatory postsynaptic currents. These structural and functional ASD-associated synaptic deficits were prevented by chronic zinc supplementation and further support zinc supplementation as a therapeutic target in ASD.

scholarly journals Next-Generation Sequencing and In Vitro Expression Study of ADAMTS13 Single Nucleotide Variants in Deep Vein Thrombosis

PLoS ONE ◽  
2016 ◽  
Vol 11 (11) ◽  
pp. e0165665 ◽  
Author(s):  
Maria Teresa Pagliari ◽  
Luca A. Lotta ◽  
Hugoline G. de Haan ◽  
Carla Valsecchi ◽  
Gloria Casoli ◽  
...  
Keyword(s):  
Deep Vein Thrombosis ◽  
Next Generation Sequencing ◽  
Single Nucleotide Variants ◽  
Expression Study ◽  
Single Nucleotide ◽  
In Vitro Expression ◽  
Vein Thrombosis ◽  
Deep Vein ◽  
Generation Sequencing

scholarly journals Co-variation of viral recombination with single nucleotide variants during virus evolution revealed by CoVaMa

2021 ◽  
Author(s):  
Shiyi Wang ◽  
Stephanea L. Sotcheff ◽  
Christian M. Gallardo ◽  
Elizabeth Jaworski ◽  
Bruce E. Torbett ◽  
...  
Keyword(s):  
Viral Evolution ◽  
Flock House Virus ◽  
Sequencing Data ◽  
Single Nucleotide Variants ◽  
Single Nucleotide ◽  
Viral Recombination ◽  
Genomic Deletions ◽  
Bioinformatic Tools ◽  
Long Read

AbstractAdaptation of viruses to their environments occurs through the acquisition of both novel Single-Nucleotide Variants (SNV) and recombination events including insertions, deletions, and duplications. The co-occurrence of SNVs in individual viral genomes during their evolution has been well-described. However, unlike covariation of SNVs, studying the correlation between recombination events with each other or with SNVs has been hampered by their inherent genetic complexity and a lack of bioinformatic tools. Here, we expanded our previously reported CoVaMa pipeline (v0.1) to measure linkage disequilibrium between recombination events and SNVs within both short-read and long-read sequencing datasets. We demonstrate this approach using long-read nanopore sequencing data acquired from Flock House virus (FHV) serially passaged in vitro. We found SNVs that were either correlated or anti-correlated with large genomic deletions generated by nonhomologous recombination that give rise to Defective-RNAs. We also analyzed NGS data from longitudinal HIV samples derived from a patient undergoing antiretroviral therapy who proceeded to virological failure. We found correlations between insertions in the p6Gag and mutations in Gag cleavage sites. This report confirms previous findings and provides insights on novel associations between SNVs and specific recombination events within the viral genome and their role in viral evolution.

scholarly journals Defining the Yeast Resistome through in vitro Evolution and Whole Genome Sequencing

2021 ◽  
Author(s):  
Sabine Ottilie ◽  
Madeline R. Luth ◽  
Erich Hellemann ◽  
Gregory M. Goldgof ◽  
Eddy Vigil ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Transcription Factors ◽  
Enrichment Analysis ◽  
Neutral Evolution ◽  
Whole Genome ◽  
Genetic Determinants ◽  
Single Nucleotide Variants ◽  
Whole Genome Analysis ◽  
Single Nucleotide

SummaryIn vitro evolution and whole genome analysis were used to comprehensively identify the genetic determinants of chemical resistance in the model microbe, Saccharomyces cerevisiae. Analysis of 355 curated, laboratory-evolved clones, resistant to 80 different compounds, demonstrates differences in the types of mutations that are identified in selected versus neutral evolution and reveals numerous new, compound-target interactions. Through enrichment analysis we further identify a set of 137 genes strongly associated with or conferring drug resistance as indicated by CRISPR-Cas9 engineering. The set of 25 most frequently mutated genes was enriched for transcription factors and for almost 25 percent of the compounds, resistance was mediated by one of 100 independently derived, gain-of-function, single nucleotide variants found in 170-amino-acid domains in two Zn2C6 transcription factors, YRR1 and YRM1 (p < 1x 10 −100). This remarkable enrichment for transcription factors as drug resistance genes may explain why it is challenging to develop effective antifungal killing agents and highlights their important role in evolution.

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