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Tuesday, April 21, 2020

DNA Methylation at the Schizophrenia and Intelligence GWAS-Implicated MIR137HG Locus May Be Associated with Disease and Cognitive Functions

The largest genome-wide association studies have identified schizophrenia and intelligence associated variants in the MIR137HG locus containing genes encoding microRNA-137 and microRNA-2682. In the present study, we investigated DNA methylation in the MIR137HG intragenic CpG island (CGI) in the peripheral blood of 44 patients with schizophrenia and 50 healthy controls. The CGI included the entire MIR137 gene and the region adjacent to the 5′-end of MIR2682. The aim of the study was to examine the relationship of the CGI methylation with schizophrenia and cognitive functioning. The methylation level of 91 CpG located in the selected region was established for each…

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Tuesday, April 21, 2020

DNA methylation analysis.

DNA methylation is a process by which methyl groups are added to cytosine or adenine. DNA methylation can change the activity of the DNA molecule without changing the sequence. Methylation of 5-methylcytosine (5mC) is widespread in both eukaryotes and prokaryotes, and it is a very important epigenetic modification event, which can regulate gene activity and influence a number of key processes such as genomic imprinting, cell differentiation, transcriptional regulation, and chromatin remodeling. Profiling DNA methylation across the genome is critical to understanding the influence of methylation in normal biology and diseases including cancer. Recent discoveries of 5-methylcytosine (5mC) oxidation derivatives…

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Sunday, September 22, 2019

Methylation of the reelin gene promoter in peripheral blood and its relationship with the cognitive function of schizophrenia patients.

There is a decrease in the expression of the reelin gene (RELN) in the brain of schizophrenia patients, which can underlie observed cognitive abnormalities. It is suggested that this decrease is caused by the hypermethylation of the RELN promoter. The aim of the study was to investigate methylation of the RELN promoter in the peripheral blood of schizophrenia patients and its association with their cognitive deficits. A modified SMRT-BS (single-molecule real-time bisulfite sequencing) was used. We determined the methylation rate of 170 CpG sites within a 1465 bp DNA region containing the entire CpG island in the RELN promoter in…

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Sunday, September 22, 2019

Relationship between Alzheimer’s disease-associated SNPs within the CLU gene, local DNA methylation and episodic verbal memory in healthy and schizophrenia subjects.

Genetic variation may impact on local DNA methylation patterns. Therefore, information about allele-specific DNA methylation (ASM) within disease-related loci has been proposed to be useful for the interpretation of GWAS results. To explore mechanisms that may underlie associations between Alzheimer’s disease (AD) and schizophrenia risk CLU gene and verbal memory, one of the most affected cognitive domains in both conditions, we studied DNA methylation in a region between AD-associated SNPs rs9331888 and rs9331896 in 72 healthy individuals and 73 schizophrenia patients. Using single-molecule real-time bisulfite sequencing we assessed the haplotype-dependent ASM in this region. We then investigated whether its methylation…

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Friday, July 19, 2019

Quantitative and multiplexed DNA methylation analysis using long-read single-molecule real-time bisulfite sequencing (SMRT-BS).

DNA methylation has essential roles in transcriptional regulation, imprinting, X chromosome inactivation and other cellular processes, and aberrant CpG methylation is directly involved in the pathogenesis of human imprinting disorders and many cancers. To address the need for a quantitative and highly multiplexed bisulfite sequencing method with long read lengths for targeted CpG methylation analysis, we developed single-molecule real-time bisulfite sequencing (SMRT-BS).Optimized bisulfite conversion and PCR conditions enabled the amplification of DNA fragments up to ~1.5 kb, and subjecting overlapping 625-1491 bp amplicons to SMRT-BS indicated high reproducibility across all amplicon lengths (r?=?0.972) and low standard deviations (=0.10) between individual CpG sites…

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Friday, July 19, 2019

CGGBP1 mitigates cytosine methylation at repetitive DNA sequences.

CGGBP1 is a repetitive DNA-binding transcription regulator with target sites at CpG-rich sequences such as CGG repeats and Alu-SINEs and L1-LINEs. The role of CGGBP1 as a possible mediator of CpG methylation however remains unknown. At CpG-rich sequences cytosine methylation is a major mechanism of transcriptional repression. Concordantly, gene-rich regions typically carry lower levels of CpG methylation than the repetitive elements. It is well known that at interspersed repeats Alu-SINEs and L1-LINEs high levels of CpG methylation constitute a transcriptional silencing and retrotransposon inactivating mechanism.Here, we have studied genome-wide CpG methylation with or without CGGBP1-depletion. By high throughput sequencing of…

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Friday, July 19, 2019

Comparative DNA methylation and gene expression analysis identifies novel genes for structural congenital heart diseases.

For the majority of congenital heart diseases (CHDs), the full complexity of the causative molecular network, which is driven by genetic, epigenetic, and environmental factors, is yet to be elucidated. Epigenetic alterations are suggested to play a pivotal role in modulating the phenotypic expression of CHDs and their clinical course during life. Candidate approaches implied that DNA methylation might have a developmental role in CHD and contributes to the long-term progress of non-structural cardiac diseases. The aim of the present study is to define the postnatal epigenome of two common cardiac malformations, representing epigenetic memory, and adaption to hemodynamic alterations,…

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Friday, July 19, 2019

Prediction of smoking by multiplex bisulfite PCR with long amplicons considering allele-specific effects on DNA methylation.

Methylation of DNA is associated with a variety of biological processes. With whole-genome studies of DNA methylation, it became possible to determine a set of genomic sites where DNA methylation is associated with a specific phenotype. A method is needed that allows detailed follow-up studies of the sites, including taking into account genetic information. Bisulfite PCR is a natural choice for this kind of task, but multiplexing is one of the most important problems impeding its implementation. To address this task, we took advantage of a recently published method based on Pacbio sequencing of long bisulfite PCR products (single-molecule real-time…

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Sunday, July 7, 2019

DNA methylation profiling using long-read Single Molecule Real-Time bisulfite sequencing (SMRT-BS).

For the past two decades, bisulfite sequencing has been a widely used method for quantitative CpG methylation detection of genomic DNA. Coupled with PCR amplicon cloning, bisulfite Sanger sequencing allows for allele-specific CpG methylation assessment; however, its time-consuming protocol and inability to multiplex has recently been overcome by next-generation bisulfite sequencing techniques. Although high-throughput sequencing platforms have enabled greater accuracy in CpG methylation quantitation as a result of increased bisulfite sequencing depth, most common sequencing platforms generate reads that are similar in length to the typical bisulfite PCR size range (~300-500 bp). Using the Pacific Biosciences (PacBio) sequencing platform, we developed…

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