A methylation-specific dot blot assay for improving specificity and sensitivity of methylation-specific PCR on DNA methylation analysis

2015 ◽  
Vol 20 (4) ◽  
pp. 839-845 ◽  
Author(s):  
Vo Thi Thuong Lan ◽  
Nguyen Thu Trang ◽  
Doan Thi Hong Van ◽  
Ta Bich Thuan ◽  
Ta Van To ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Methylation Analysis ◽  
Dot Blot ◽  
Methylation Specific Pcr ◽  
Specific Pcr ◽  
Specificity And Sensitivity ◽  
Dot Blot Assay ◽  
Dna Methylation Analysis

scholarly journals A systematic comparison of quantitative high-resolution DNA methylation analysis and methylation-specific PCR

Epigenetics ◽  
2012 ◽  
Vol 7 (7) ◽  
pp. 772-780 ◽  
Author(s):  
Rainer Claus ◽  
Stefan Wilop ◽  
Thomas Hielscher ◽  
Miriam Sonnet ◽  
Edgar Dahl ◽  
...  
Keyword(s):  
Dna Methylation ◽  
High Resolution ◽  
Methylation Analysis ◽  
Systematic Comparison ◽  
Methylation Specific Pcr ◽  
Specific Pcr ◽  
Dna Methylation Analysis

scholarly journals Multiplexed DNA Methylation Analysis in Colorectal Cancer Using Liquid Biopsy and Its Diagnostic and Predictive Value

2021 ◽  
Vol 43 (3) ◽  
pp. 1419-1435
Author(s):  
Walter Pulverer ◽  
Kristi Kruusmaa ◽  
Silvia Schönthaler ◽  
Jasmin Huber ◽  
Marko Bitenc ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Dna Methylation ◽  
Therapy Monitoring ◽  
Methylation Status ◽  
Methylation Analysis ◽  
Microarray Experiments ◽  
Specific Pcr ◽  
Colonic Adenomas ◽  
Methylation Sensitive Restriction Enzyme ◽  
Dna Methylation Analysis

Early diagnosis of colorectal cancer (CRC) is of high importance as prognosis depends on tumour stage at the time of diagnosis. Detection of tumour-specific DNA methylation marks in cfDNA has several advantages over other approaches and has great potential for solving diagnostic needs. We report here the identification of DNA methylation biomarkers for CRC and give insights in our methylation-sensitive restriction enzyme coupled qPCR (MSRE-qPCR) system. Targeted microarrays were used to investigate the DNA methylation status of 360 cancer-associated genes. Validation was done by qPCR-based approaches. A focus was on investigating marker performance in cfDNA from 88 patients (44 CRC, 44 controls). Finally, the workflow was scaled-up to perform 180plex analysis on 110 cfDNA samples, to identify a DNA methylation signature for advanced colonic adenomas (AA). A DNA methylation signature (n = 44) was deduced from microarray experiments and confirmed by quantitative methylation-specific PCR (qMSP) and by MSRE-qPCR, providing for six genes’ single areas under the curve (AUC) values of >0.85 (WT1, PENK, SPARC, GDNF, TMEFF2, DCC). A subset of the signatures can be used for patient stratification and therapy monitoring for progressed CRC with liver metastasis using cfDNA. Furthermore, we identified a 35-plex classifier for the identification of AAs with an AUC of 0.80.

scholarly journals CADM1, MAL and miR124-2 methylation analysis in cervical scrapes to detect cervical and endometrial cancer

2014 ◽  
Vol 67 (12) ◽  
pp. 1067-1071 ◽  
Author(s):  
Lise M A De Strooper ◽  
Marjolein van Zummeren ◽  
Renske D M Steenbergen ◽  
Maaike C G Bleeker ◽  
Albertus T Hesselink ◽  
...  
Keyword(s):  
Cervical Cancer ◽  
Dna Methylation ◽  
Endometrial Cancer ◽  
Gene Promoter ◽  
Intraepithelial Neoplasia ◽  
Promoter Hypermethylation ◽  
Methylation Analysis ◽  
Cervical Intraepithelial Neoplasia Grade ◽  
Specific Pcr ◽  
Dna Methylation Analysis

AimsGene promoter hypermethylation is recognised as an essential early step in carcinogenesis, indicating important application areas for DNA methylation analysis in early cancer detection. The current study was set out to assess the performance of CADM1, MAL and miR124-2 methylation analysis in cervical scrapes for detection of cervical and endometrial cancer.MethodsA series of cervical scrapes of women with cervical (n=79) or endometrial (n=21) cancer, cervical intraepithelial neoplasia grade 3 (CIN3) (n=16) or CIN2 (n=32), and women without evidence of CIN2 or worse (n=120) were assessed for methylation of CADM1, MAL and miR124-2. Methylation analysis was done by the PreCursor-M assay, a multiplex quantitative methylation-specific PCR.ResultsAll samples of women with cervical cancer (79/79, 100%), independent of the histotype, and 76% (16/21; 95% CI 58.0% to 94.4%) of women with endometrial cancer scored positive for DNA methylation for at least one of the three genes. In women without cancer, methylation frequencies increased significantly with severity of disease from 19.2% (23/120; 95% CI 12.1% to 26.2%) in women without CIN2 or worse to 37.5% (12/32; 95% CI 20.7% to 54.3%) and 68.8% (11/16; 95% CI 46.0% to 91.5%) in women with CIN2 and CIN3, respectively. Overall methylation positivity and the number of methylated genes increased proportionally to the lesion severity.ConclusionsDNA methylation analysis of CADM1, MAL and miR124-2 in cervical scrapes consistently detects cervical cancer and the majority of CIN3 lesions, and has the capacity to broaden its use on cervical scrapes through the detection of a substantial subset of endometrial carcinomas.

scholarly journals P46.03 DNA Methylation Analysis in Smokers and Non-Smokers (Passive and ex-Smokers)

2021 ◽  
Vol 16 (3) ◽  
pp. S490
Author(s):  
D.M. Aguilar-Beltrán ◽  
A.G. Alcázar-Ramos ◽  
A.L. Vega-Rodríguez ◽  
D.G. García-Gutiérrez ◽  
A.D. Bertadillo-Jilote ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Methylation Analysis ◽  
Dna Methylation Analysis

scholarly journals DNA Methylation in Solid Tumors: Functions and Methods of Detection

2021 ◽  
Vol 22 (8) ◽  
pp. 4247
Author(s):  
Andrea Martisova ◽  
Jitka Holcakova ◽  
Nasim Izadi ◽  
Ravery Sebuyoya ◽  
Roman Hrstka ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Solid Tumors ◽  
Epigenetic Modification ◽  
Single Gene ◽  
Restriction Enzymes ◽  
Methylation Analysis ◽  
Cpg Dinucleotides ◽  
Sequencing Technologies ◽  
Genome Level ◽  
Dna Methylation Analysis

DNA methylation, i.e., addition of methyl group to 5′-carbon of cytosine residues in CpG dinucleotides, is an important epigenetic modification regulating gene expression, and thus implied in many cellular processes. Deregulation of DNA methylation is strongly associated with onset of various diseases, including cancer. Here, we review how DNA methylation affects carcinogenesis process and give examples of solid tumors where aberrant DNA methylation is often present. We explain principles of methods developed for DNA methylation analysis at both single gene and whole genome level, based on (i) sodium bisulfite conversion, (ii) methylation-sensitive restriction enzymes, and (iii) interactions of 5-methylcytosine (5mC) with methyl-binding proteins or antibodies against 5mC. In addition to standard methods, we describe recent advances in next generation sequencing technologies applied to DNA methylation analysis, as well as in development of biosensors that represent their cheaper and faster alternatives. Most importantly, we highlight not only advantages, but also disadvantages and challenges of each method.

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