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Epigenerics or methylation or hypermethylation or hypomethylation or HIV

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https://www.readbyqxmd.com/read/29224242/dermoscopic-observations-in-disseminated-cryptococcosis-with-cutaneous-involvement
#1
M Sławińska, M Hlebowicz, E Iżycka-Świeszewska, M Sikorska, M Sokołowska-Wojdyło, T Smiatacz, R Nowicki, M Sobjanek
A 26-year-old female patient with a history of intravenous drug abuse, diagnosed with acquired immunodeficiency syndrome (AIDS) 6 months before. AIDS defining disease was Pneumocystis jiroveci pneumonia. The patient presented with severely decreased CD4 cell count of 7 cells/mm3 and relatively low HIV viral load of 281 copies/mL. Shortly after initiation of antiretroviral therapy (ART), was diagnosed with disseminated cryptococcosis with central nervous system and skin involvement. This article is protected by copyright...
December 10, 2017: Journal of the European Academy of Dermatology and Venereology: JEADV
https://www.readbyqxmd.com/read/29224182/stable-and-rigid-dtpa-like-paramagnetic-tags-suitable-for-in-vitro-and-in-situ-protein-nmr-analysis
#2
Jia-Liang Chen, Yu Zhao, Yan-Jun Gong, Bin-Bin Pan, Xiao Wang, Xun-Cheng Su
Organic synthesis of a ligand with high binding affinities for paramagnetic lanthanide ions is an effective way of generating paramagnetic effects on proteins. These paramagnetic effects manifested in high-resolution NMR spectroscopy are valuable dynamic and structural restraints of proteins and protein-ligand complexes. A paramagnetic tag generally contains a metal chelating moiety and a reactive group for protein modification. Herein we report two new DTPA-like tags, 4PS-PyDTTA and 4PS-6M-PyDTTA that can be site-specifically attached to a protein with a stable thioether bond...
December 9, 2017: Journal of Biomolecular NMR
https://www.readbyqxmd.com/read/29224170/affinity-based-enrichment-techniques-for-the-genome-wide-analysis-of-5-hydroxymethylcytosine
#3
John P Thomson, Richard R Meehan
Since its initial characterization in 2009 there has been a great degree of interest in comparative profiling of 5-hydroxymethylcytosine (5hmC) nucleotides in vertebrate DNA. Through a host of genome-wide studies the distribution of 5hmC has been mapped in a range of cell lines, tissue types and organisms; the majority of which have been generated through affinity-based methods for 5hmC enrichment. Although recent advances in the field have resulted in the ability to investigate the levels of both methylated and hydroxymethylated cytosines at single base resolution, such studies are still relatively cost-prohibitive as well as technically challenging...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224169/multiplexing-for-oxidative-bisulfite-sequencing-oxbs-seq
#4
Kristina Kirschner, Felix Krueger, Anthony R Green, Tamir Chandra
DNA modifications, especially methylation, are known to play a crucial part in many regulatory processes in the cell. Recently, 5-hydroxymethylcytosine (5hmC) was discovered, a DNA modification derived as an intermediate of 5-methylcytosine (5mC) oxidation. Efforts to gain insights into function of this DNA modification are underway and several methods were recently described to assess 5hmC levels using sequencing approaches. Here we integrate adaptation based multiplexing and high-efficiency library prep into the oxidative Bisulfite Sequencing (oxBS-seq) workflow reducing the starting amount and cost per sample to identify 5hmC levels genome-wide...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224167/dna-methylation-analysis-of-free-circulating-dna-in-body-fluids
#5
Maria Jung, Glen Kristiansen, Dimo Dietrich
Circulating cell-free DNA in body fluids is an analyte of great interest in basic and clinical research. The analyses of DNA methylation and hydroxymethylation patterns in body fluids might allow one to determine the certain state of a disease, in particular of cancer. DNA methylation biomarkers in liquid biopsies, i.e. blood plasma samples, may help optimizing personalized therapy for individual patients. DNA methylation analyses of specific loci usually require a bisulfite conversion of the DNA, which requires a sufficiently high amount of DNA at the appropriate concentration...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224166/dna-methylation-analysis-from-blood-spots-increasing-yield-and-quality-for-genome-wide-and-locus-specific-methylation-analysis
#6
Akram Ghantous, Hector Hernandez-Vargas, Zdenko Herceg
Blood represents an easily accessible human tissue for numerous research and clinical applications, including surrogate roles for biomarkers of other tissues. Dried blood spots (DBS) are space- and cost-efficient storage forms of blood while stably retaining many of its chemical constituents. Consequently, neonatal DBS are routinely collected in many countries, and their biobanks represent gold mines for research. However, the utility of DBS is restricted by the limited amount and quality of extractable biomolecules (including DNA), especially for genome-wide profiling...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224165/helper-dependent-chain-reaction-hdcr-for-selective-amplification-of-methylated-dna-sequences
#7
Susan M Mitchell, Keith N Rand, Zheng-Zhou Xu, Thu Ho, Glenn S Brown, Jason P Ross, Peter L Molloy
Over the last few years a number of restriction enzymes that cut DNA only if cytosines within their recognition sequences are methylated have been characterized and become commercially available. Cleavage with these enzymes to release DNA fragments in a methylation-dependent manner can be combined with a novel method of amplification, Helper Dependent Chain Reaction (HDCR), to selectively amplify these fragments. HDCR uses "Helper" oligonucleotides as templates for extension of the free 3' end of target fragments to incorporate tag sequences at the ends of fragments...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224164/hairpin-bisulfite-sequencing-synchronous-methylation-analysis-on-complementary-dna-strands-of-individual-chromosomes
#8
Pascal Giehr, Jörn Walter
The accurate and quantitative detection of 5-methylcytosine is of great importance in the field of epigenetics. The method of choice is usually bisulfite sequencing because of the high resolution and the possibility to combine it with next generation sequencing. Nevertheless, also this method has its limitations. Following the bisulfite treatment DNA strands are no longer complementary such that in a subsequent PCR amplification the DNA methylation patterns information of only one of the two DNA strand is preserved...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224163/methylation-sensitive-high-resolution-melting-ms-hrm
#9
Dianna Hussmann, Lise Lotte Hansen
Methylation-Sensitive High Resolution Melting (MS-HRM) is an in-tube, PCR-based method to detect methylation levels at specific loci of interest. A unique primer design facilitates a high sensitivity of the assays enabling detection of down to 0.1-1% methylated alleles in an unmethylated background.Primers for MS-HRM assays are designed to be complementary to the methylated allele, and a specific annealing temperature enables these primers to anneal both to the methylated and the unmethylated alleles thereby increasing the sensitivity of the assays...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224162/methylation-specific-multiplex-ligation-dependent-probe-amplification-ms-mlpa
#10
Cathy B Moelans, Lilit Atanesyan, Suvi P Savola, Paul J van Diest
This chapter describes a method for the rapid assessment of promoter hypermethylation levels or methylation of imprinted regions in human genomic DNA extracted from various sources using methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA). Multiplex ligation-dependent probe amplification (MLPA) is a powerful and easy-to-perform PCR-based technique that can identify gains, amplifications, losses, deletions, methylation and mutations of up to 55 targets in a single reaction, while requiring only minute quantities of DNA (about 50 ng) extracted from blood, fresh frozen or formalin-fixed paraffin-embedded materials...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224161/quantitative-region-specific-dna-methylation-analysis-by-the-epityper%C3%A2-technology
#11
Sonja Kunze
DNA methylation plays a profound role in development and health as well as development and progression of disease. High-throughput quantitative DNA methylation analysis is therefore crucial for the study of the normal physiology of the epigenome and its dysregulation in disease. Many target areas are identified by a range of emerging genome-wide cytosine methylation techniques, but these whole genome scans usually only provide methylation data for a few individual CpG sites (CpGs) within a region. The EpiTYPER™ assay is a region-specific method for the detection and quantitative analysis of DNA methylation that allows performing a high-resolution scan of selected regions...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224160/methylight-and-digital-methylight
#12
Mihaela Campan, Daniel J Weisenberger, Binh Trinh, Peter W Laird
MethyLight is a quantitative, fluorescence-based, real-time PCR method to sensitively detect and quantify DNA methylation of candidate regions of the genome. MethyLight is uniquely suited for detecting low-frequency methylated DNA regions against a high background of unmethylated DNA, as it combines methylation-specific priming with methylation-specific fluorescent probing. The quantitative accuracy of real-time PCR and the ability to design bisulfite-dependent, DNA methylation-independent control reactions together allow for a quantitative assessment of these low frequency methylation events...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224159/quantitation-of-dna-methylation-by-quantitative-multiplex-methylation-specific-pcr-qm-msp-assay
#13
Mary Jo Fackler, Saraswati Sukumar
The defining feature of the Quantitative Multiplex Methylation-Specific PCR (QM-MSP) method to sensitively quantify DNA methylation is the two-step PCR approach for a multiplexed analysis of a panel of up to 12 genes in clinical samples with minimal quantities of DNA. In the first step, for up to 12 genes tested, one pair of gene-specific primers (forward and reverse) amplifies the methylated and unmethylated copies of the same gene simultaneously and in multiplex, in one PCR reaction. This methylation-independent amplification step produces amplicons of up to 109 copies per μL after 36 cycles of PCR...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224158/methylation-specific-pcr
#14
João Ramalho-Carvalho, Rui Henrique, Carmen Jerónimo
Cytosine methylation is a DNA modification generally associated with transcriptional silencing. Factors that regulate methylation have been linked to human disease, yet how they contribute to malignancies remains largely unknown. Methylation of DNA can change the functional state of regulatory regions, but does not change the Watson-Crick base pairing of cytosine. Moreover, sequence symmetry of CpGs enables propagation of the methyl mark through cell division. This potential for inheritance coupled with the fact that DNA methylation patterns change during development and disease partially explains the interest in DNA methylation as a memory module...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224157/quantitative-dna-methylation-analysis-at-single-nucleotide-resolution-by-pyrosequencing%C3%A2
#15
Florence Busato, Emelyne Dejeux, Hafida El Abdalaoui, Ivo Glynne Gut, Jörg Tost
Many protocols for gene-specific DNA methylation analysis are either labor intensive, not quantitative and/or limited to the measurement of the methylation status of only one or very few CpG positions. Pyrosequencing is a real-time sequencing technology that overcomes these limitations. After bisulfite modification of genomic DNA, a region of interest is amplified by PCR with one of the two primers being biotinylated. The PCR generated template is rendered single-stranded and a pyrosequencing primer is annealed to analyze quantitatively cytosine methylation...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224156/multiplexed-and-sensitive-dna-methylation-testing-using-methylation-sensitive-restriction-enzymes-msre-qpcr
#16
Gabriel Beikircher, Walter Pulverer, Manuela Hofner, Christa Noehammer, Andreas Weinhaeusel
DNA methylation is a chemically stable key-player in epigenetics. In the vertebrate genome the 5-methyl cytosine (5mC) has been found almost exclusively in the CpG dinucleotide context. CpG dinucleotides are enriched in CpG islands very frequently located within or close to gene promoters. Analyses of DNA methylation changes in human diagnostics have been conducted classically using methylation-sensitive restriction enzymes (MSRE). Since the discovery of bisulfite conversion-based sequencing and PCR assays, MSRE-based PCR assays have been less frequently used, although especially in the field of cancer epigenetics MSRE-based genome-wide discovery and targeted screening applications have been and are still performed successfully...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224155/targeted-bisulfite-sequencing-using-the-seqcap-epi-enrichment-system
#17
Jennifer Wendt, Heidi Rosenbaum, Todd A Richmond, Jeffrey A Jeddeloh, Daniel L Burgess
Cytosine methylation has been shown to have a role in a host of biological processes. In mammalian biology these include stem cell differentiation, embryonic development, genomic imprinting, inflammation, and silencing of transposable elements. Given the central importance of these processes, it is not surprising to find aberrant cytosine methylation patterns associated with many disorders in humans, including cancer, cardiovascular disease, and neurological disease. While whole genome shotgun bisulfite sequencing (WGBS) has recently become feasible, generating high sequence coverage data for the entire genome is expensive, both in terms of money and analysis time, when generally only a small subset of the genome is of interest to most researchers...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224154/large-scale-targeted-dna-methylation-analysis-using-bisulfite-padlock-probes
#18
Dinh Diep, Nongluk Plongthongkum, Kun Zhang
Bisulfite padlock probes (BSPP) are a method for the targeted quantification of DNA methylation in mammalian genomes. They can simultaneously characterize the level of methylcytosine modification in a large number of targeted regions at single-base resolution. A major advantage of BSPP is that it allows the flexible capture of an arbitrary subset of genomic regions (hundreds to hundreds of thousands of genomic loci) in single-tube reactions. Large number of samples can be processed efficiently and converted into multiplexed sequencing libraries with only three enzymatic steps, without the conventional library preparation procedures...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224153/multiplexed-dna-methylation-analysis-of-target-regions-using-microfluidics-fluidigm
#19
Martyna Adamowicz, Klio Maratou, Timothy J Aitman
Whole genome shotgun bisulfite sequencing is a method used to generate genome-wide methylation profiles. There are many available protocols to validate the results of this genome-wide method, but they mostly share the limitation of measuring methylation at a small number of CpG positions in small numbers of samples. We developed a multiplexed DNA methylation analysis protocol, which allows for the simultaneous quantitative measurement of cytosine methylation at single nucleotide resolution in 48 PCR amplicons and 48 samples utilizing the microfluidic system established by Fluidigm...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29224152/microdroplet-pcr-for-highly-multiplexed-targeted-bisulfite-sequencing
#20
H Kiyomi Komori, Sarah A LaMere, Traver Hart, Steven R Head, Ali Torkamani, Daniel R Salomon
Many methods exist for examining CpG DNA methylation. However, many of these are qualitative, laborious to apply to a large number of genes simultaneously, or are not easy to target to specific regions of interest. Microdroplet PCR-based bisulfite sequencing allows for quantitative single base resolution analysis of investigator selected regions of interest. Following bisulfite conversion of genomic DNA, targeted microdroplet PCR is conducted with custom primer libraries. Samples are then fragmented, concatenated, and sequenced by high-throughput sequencing...
2018: Methods in Molecular Biology
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