Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV Detection
Alterations in global DNA methylation are implicated in various pathophysiological processes. The development of simple and quick, yet robust, methods to assess DNA methylation is required to facilitate its measurement and interpretation in clinical practice. We describe a highly sensitive and repro...
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| Format: | Article |
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Wiley
2017-01-01
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| Series: | Journal of Analytical Methods in Chemistry |
| Online Access: | http://dx.doi.org/10.1155/2017/4065892 |
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| author | Angelo Zinellu Elisabetta Sotgiu Stefano Assaretti Salvatore Sotgia Panagiotis Paliogiannis Gianfranco Pintus Arduino A. Mangoni Ciriaco Carru |
| author_facet | Angelo Zinellu Elisabetta Sotgiu Stefano Assaretti Salvatore Sotgia Panagiotis Paliogiannis Gianfranco Pintus Arduino A. Mangoni Ciriaco Carru |
| author_sort | Angelo Zinellu |
| collection | DOAJ |
| description | Alterations in global DNA methylation are implicated in various pathophysiological processes. The development of simple and quick, yet robust, methods to assess DNA methylation is required to facilitate its measurement and interpretation in clinical practice. We describe a highly sensitive and reproducible capillary electrophoresis method with UV detection for the separation and detection of cytosine and methylcytosine, after formic acid hydrolysis of DNA extracted from human whole blood. Hydrolysed samples were dried and resuspended with water and directly injected into the capillary without sample derivatization procedures. The use of a run buffer containing 50 mmol/L BIS-TRIS propane (BTP) phosphate buffer at pH 3.25 and 60 mmol/L sodium acetate buffer at pH 3.60 (4 : 1, v/v) allowed full analyte identification within 11 min. Precision tests indicated an elevated reproducibility with an interassay CV of 1.98% when starting from 2 μg of the extracted DNA. The method was successfully tested by measuring the DNA methylation degree both in healthy volunteers and in reference calf thymus DNA. |
| format | Article |
| id | doaj-art-1cd89d60def6461a9ccefe9e40a3bf2c |
| institution | OA Journals |
| issn | 2090-8865 2090-8873 |
| language | English |
| publishDate | 2017-01-01 |
| publisher | Wiley |
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| series | Journal of Analytical Methods in Chemistry |
| spelling | doaj-art-1cd89d60def6461a9ccefe9e40a3bf2c2025-08-20T02:01:46ZengWileyJournal of Analytical Methods in Chemistry2090-88652090-88732017-01-01201710.1155/2017/40658924065892Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV DetectionAngelo Zinellu0Elisabetta Sotgiu1Stefano Assaretti2Salvatore Sotgia3Panagiotis Paliogiannis4Gianfranco Pintus5Arduino A. Mangoni6Ciriaco Carru7Department of Biomedical Sciences, University of Sassari, Sassari, ItalyDepartment of Biomedical Sciences, University of Sassari, Sassari, ItalyDepartment of Biomedical Sciences, University of Sassari, Sassari, ItalyDepartment of Biomedical Sciences, University of Sassari, Sassari, ItalyDepartment of Clinical and Experimental Medicine, University of Sassari, Sassari, ItalyDepartment of Biomedical Sciences, College of Health Sciences, Qatar University, Doha 2713, QatarDepartment of Clinical Pharmacology, College of Medicine and Public Health, Flinders University, Adelaide, SA, AustraliaDepartment of Biomedical Sciences, University of Sassari, Sassari, ItalyAlterations in global DNA methylation are implicated in various pathophysiological processes. The development of simple and quick, yet robust, methods to assess DNA methylation is required to facilitate its measurement and interpretation in clinical practice. We describe a highly sensitive and reproducible capillary electrophoresis method with UV detection for the separation and detection of cytosine and methylcytosine, after formic acid hydrolysis of DNA extracted from human whole blood. Hydrolysed samples were dried and resuspended with water and directly injected into the capillary without sample derivatization procedures. The use of a run buffer containing 50 mmol/L BIS-TRIS propane (BTP) phosphate buffer at pH 3.25 and 60 mmol/L sodium acetate buffer at pH 3.60 (4 : 1, v/v) allowed full analyte identification within 11 min. Precision tests indicated an elevated reproducibility with an interassay CV of 1.98% when starting from 2 μg of the extracted DNA. The method was successfully tested by measuring the DNA methylation degree both in healthy volunteers and in reference calf thymus DNA.http://dx.doi.org/10.1155/2017/4065892 |
| spellingShingle | Angelo Zinellu Elisabetta Sotgiu Stefano Assaretti Salvatore Sotgia Panagiotis Paliogiannis Gianfranco Pintus Arduino A. Mangoni Ciriaco Carru Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV Detection Journal of Analytical Methods in Chemistry |
| title | Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV Detection |
| title_full | Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV Detection |
| title_fullStr | Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV Detection |
| title_full_unstemmed | Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV Detection |
| title_short | Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV Detection |
| title_sort | evaluation of global genomic dna methylation in human whole blood by capillary electrophoresis uv detection |
| url | http://dx.doi.org/10.1155/2017/4065892 |
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