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. 2020 Aug 3;10:13026. doi: 10.1038/s41598-020-69750-0

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Methodology of DNA methylation analysis for DBS. Step 1: Collect blood and fill the spots on filter papers; Step 2: Extract the DNA from each Guthrie cards and from Whole Blood; Step 3: Bisulfite treatment, converting unmethylated cytosine to uracil; Step 4: PCR amplification; Step 5: Allele discrimination through HRM methodology: Step 6: Results analysis and diagnostic.*It does not distinguish between the different molecular causes related to PWS and AS.