Proceedings of the Conference on Natural Resources And Life Sciences 2022 (NRLS-BIO 2022)

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Methylation Specific PCR (MSP): Nested PCR vs Unnested PCR

Authors
Farizky Martriano Humardani1, 2, Lisa Thalia Mulyanata1, Lady Theresa Adeodata Tanaya1, Risma Ikawaty1, Heru Wijono1, Hikmawan Wahyu Sulistomo2, Dini Kesuma3, Sulistyo Emantoko Dwi Putra4, *
1Faculty of Medicine, University of Surabaya, Surabaya, 60292, Indonesia
2Department of Biomedical Science, Faculty of Medicine, Universitas Brawijaya, Malang, 65112, Indonesia
3Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Surabaya, Surabaya, 60292, Indonesia
4Faculty of Biotechnology, University of Surabaya, Surabaya, 60292, Indonesia
*Corresponding author. Email: emantoko@staff.ubaya.ac.id
Corresponding Author
Sulistyo Emantoko Dwi Putra
Available Online 30 December 2023.
DOI
10.2991/978-94-6463-322-1_12How to use a DOI?
Keywords
Bisulfite; Methylation; Nested MSP; Unnested MSP
Abstract

Methylation-specific PCR (MSP) is a valuable technique for studying DNA methylation patterns due to its straightforward design and implementation, high sensitivity in detecting methylated DNA, and ability to analyze large sample sizes cost-effectively rapidly. However, researchers need to be cautious when working with new samples or samples stored in the freezer for an extended period. Freezing does not prevent the action of DNAase enzymes, which can lead to reduced DNA extraction yields. During MSP, DNA undergoes changes, and PCR is performed using two sets of primers that specifically target methylated and unmethylated DNA regions. The use of bisulfite conversion treatment, an essential step in methylation analysis, presents significant challenges. One challenge is that bisulfite treatment can cause DNA fragmentation, particularly when the initial DNA concentration is low. In nested MSP, where an additional round of PCR is performed, smearing may occur due to the high DNA concentrations used. Therefore, for new samples with a DNA concentration of 1–10 µg/µL, it is recommended to use the unnested MSP technique prior to bisulfite conversion treatment.

Copyright
© 2023 The Author(s)
Open Access
Open Access This chapter is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International License (http://creativecommons.org/licenses/by-nc/4.0/), which permits any noncommercial 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.

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Volume Title
Proceedings of the Conference on Natural Resources And Life Sciences 2022 (NRLS-BIO 2022)
Series
Advances in Biological Sciences Research
Publication Date
30 December 2023
ISBN
10.2991/978-94-6463-322-1_12
ISSN
2468-5747
DOI
10.2991/978-94-6463-322-1_12How to use a DOI?
Copyright
© 2023 The Author(s)
Open Access
Open Access This chapter is licensed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International License (http://creativecommons.org/licenses/by-nc/4.0/), which permits any noncommercial 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.

Cite this article

risenwbib
TY  - CONF
AU  - Farizky Martriano Humardani
AU  - Lisa Thalia Mulyanata
AU  - Lady Theresa Adeodata Tanaya
AU  - Risma Ikawaty
AU  - Heru Wijono
AU  - Hikmawan Wahyu Sulistomo
AU  - Dini Kesuma
AU  - Sulistyo Emantoko Dwi Putra
PY  - 2023
DA  - 2023/12/30
TI  - Methylation Specific PCR (MSP): Nested PCR vs Unnested PCR
BT  - Proceedings of the Conference on Natural Resources And Life Sciences 2022 (NRLS-BIO 2022)
PB  - Atlantis Press
SP  - 90
EP  - 95
SN  - 2468-5747
UR  - https://doi.org/10.2991/978-94-6463-322-1_12
DO  - 10.2991/978-94-6463-322-1_12
ID  - Humardani2023
ER  -