Volume 66 Issue 4
Jun.  2022
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WANG Di, NIU Chunyan, WANG Zhidong, YANG Jiayi, DONG Lianhua, GAO Yunhua. Detection Methods and Accuracy Analysis of DNA Methylation[J]. Metrology Science and Technology, 2022, 66(4): 55-62. doi: 10.12338/j.issn.2096-9015.2021.0609
Citation: WANG Di, NIU Chunyan, WANG Zhidong, YANG Jiayi, DONG Lianhua, GAO Yunhua. Detection Methods and Accuracy Analysis of DNA Methylation[J]. Metrology Science and Technology, 2022, 66(4): 55-62. doi: 10.12338/j.issn.2096-9015.2021.0609

Detection Methods and Accuracy Analysis of DNA Methylation

doi: 10.12338/j.issn.2096-9015.2021.0609
  • Available Online: 2022-03-21
  • Publish Date: 2022-06-02
  • DNA methylation is a common epigenetic modification in eukaryotes and plays a crucial role in the evolution of many physiological and disease processes. Methylated DNA has become a type of key biomarker, and accurate quantification of the methylation levels is of great value for clinical management and basic scientific research. However, current results based on different platforms and different methods often have more significant differences in results, and this uncertainty poses a great potential for its application. In this paper, we summarize some typical detection methods for the locus-specific methylation analysis, especially highlighting the application of digital PCR in DNA methylation detection with high sensitivity and accuracy, and analyze the key factors affecting the result accuracy, and finally prospect the establishment of a standard evaluation system for quantifying methylated DNA.
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  • [1]
    Jin Z, Liu Y. DNA methylation in human diseases[J]. Genes & diseases, 2018, 5(1): 1-8.
    Jones PA, Takai D. The role of DNA methylation in mammalian epigenetics[J]. Science, 2001, 293(5532): 1068-1070. doi: 10.1126/science.1063852
    Shen L, Song CX, He C, et al. Mechanism and function of oxidative reversal of DNA and RNA methylation[J]. Annu Rev Biochem, 2014, 83: 585-614. doi: 10.1146/annurev-biochem-060713-035513
    Schubeler D. Function and information content of DNA methylation[J]. Nature, 2015, 517(7534): 321-326. doi: 10.1038/nature14192
    Gardiner-Garden M, Frommer M. CpG islands in vertebrate genomes[J]. Journal of molecular biology, 1987, 196(2): 261-282. doi: 10.1016/0022-2836(87)90689-9
    Cross SH, Charlton JA, Nan X, et al. Purification of CpG islands using a methylated DNA binding column[J]. Nature genetics, 1994, 6(3): 236-244. doi: 10.1038/ng0394-236
    Robertson K D. DNA methylation and human disease[J]. Nature reviews Genetics, 2005, 6(8): 597-610. doi: 10.1038/nrg1655
    Kulis M, Esteller M. DNA methylation and cancer[J]. Advances in genetics, 2010, 70: 27-56.
    How KA, Nielsen HM, Tost J. DNA methylation based biomarkers: practical considerations and applications[J]. Biochimie, 2012, 94(11): 2314-2337. doi: 10.1016/j.biochi.2012.07.014
    Van DVL, Speeckaert R, Van G D, et al. DNA methylation-based biomarkers in serum of patients with breast cancer[J]. Mutation research, 2012, 751(2): 304-325. doi: 10.1016/j.mrrev.2012.06.001
    Heyn H, Esteller M. DNA methylation profiling in the clinic: applications and challenges[J]. Nature reviews Genetics, 2012, 13(10): 679-692. doi: 10.1038/nrg3270
    Šestáková Š, Šálek C, Remešová H. DNA Methylation Validation Methods: a Coherent Review with Practical Comparison[J]. Biological procedures online, 2019, 21: 19-25. doi: 10.1186/s12575-019-0107-z
    王玲, 彭付端, 赵慧, 等. 焦磷酸测序和MassARRAY定量检测DNA甲基化在年龄推断中的差异[J]. 南方医科大学学报, 2020, 40(12): 145-151.
    Feng L, Lou J. DNA Methylation Analysis[J]. Methods Mol Biol, 2019, 1894: 181-227.
    Kurdyukov S, Bullock M. DNA Methylation Analysis: Choosing the Right Method[J]. Biology, 2016, 5(3): 1-22.
    Nakagawa T, Wakui M, Hayashida T, et al. Intensive optimization and evaluation of global DNA methylation quantification using LC-MS/MS[J]. Analytical and bioanalytical chemistry, 2019, 411(27): 7221-7231. doi: 10.1007/s00216-019-02115-3
    杨佳怡, 傅博强, 李曼莉, 等. 基于DNA提取效率校正的数字PCR定量方法[J]. 计量科学与技术, 2020(10): 32-36. doi: 10.3969/j.issn.2096-9015.2020.10.09
    Torres AL, Barrientos EY, Wrobel K, et al. Selective derivatization of cytosine and methylcytosine moieties with 2-bromoacetophenone for submicrogram DNA methylation analysis by reversed phase HPLC with spectrofluorimetric detection[J]. Analytical chemistry, 2011, 83(20): 7999-8005. doi: 10.1021/ac2020799
    Gowher H, Leismann O, Jeltsch A. DNA of Drosophila melanogaster contains 5-methylcytosine. The EMBO journal[J]. 2000, 19(24): 6918-6923.
    曹雷, 郭利娟, 郭晓锦, 等. 癌症液体活检新思路: 数字PCR检测DNA甲基化[J]. 生物化学与生物物理进展, 2019, 46(11): 50-65.
    Laird PW. Principles and challenges of genomewide DNA methylation analysis[J]. Nature reviews Genetics, 2010, 11(3): 191-203. doi: 10.1038/nrg2732
    Waalwijk C, Flavell RA. DNA methylation at a CCGG sequence in the large intron of the rabbit beta-globin gene: tissue-specific variations[J]. Nucleic acids research, 1978, 5(12): 4631-4634. doi: 10.1093/nar/5.12.4631
    Kaput J, Sneider TW. Methylation of somatic vs germ cell DNAs analyzed by restriction endonuclease digestions[J]. Nucleic acids research, 1979, 7(8): 2303-2322. doi: 10.1093/nar/7.8.2303
    Gautier F, Bünemann H, Grotjahn L. Analysis of calf-thymus satellite DNA: evidence for specific methylation of cytosine in C-G sequences[J]. European journal of biochemistry, 1977, 80(1): 175-183. doi: 10.1111/j.1432-1033.1977.tb11869.x
    Vander P LH, Flavell RA. DNA methylation in the human gamma delta beta-globin locus in erythroid and nonerythroiqd tissues[J]. Cell, 1980, 19(4): 947-958. doi: 10.1016/0092-8674(80)90086-0
    Hayatsu H, Wataya Y, Kazushige K. The addition of sodium bisulfite to uracil and to cytosine[J]. J Am Chem Soc, 1970, 92(3): 724-726. doi: 10.1021/ja00706a062
    Hayatsu H. Discovery of bisulfite-mediated cytosine conversion to uracil, the key reaction for DNA methylation analysis-a personal account[J]. Proceedings of the Japan Academy Series B, 2008, 84(8): 321-330. doi: 10.2183/pjab.84.321
    Herman JG, Graff JR, Myöhänen S, et al. Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands[J]. Proceedings of the National Academy of Sciences of the United States of America, 1996, 93(18): 9821-9826. doi: 10.1073/pnas.93.18.9821
    Scher MB, Elbaum MB, Mogilevkin Y, et al. Detecting DNA methylation of the BCL2, CDKN2A and NID2 genes in urine using a nested methylation specific polymerase chain reaction assay to predict bladder cancer[J]. The Journal of urology, 2012, 188(6): 2101-2107. doi: 10.1016/j.juro.2012.08.015
    Husseiny MI, Kuroda A, Kaye AN, et al. Development of a quantitative methylation-specific polymerase chain reaction method for monitoring beta cell death in type 1 diabetes[J]. PLoS One, 2012, 7(10): 47942. doi: 10.1371/journal.pone.0047942
    Ronaghi M, Uhlén M, Nyrén P. A sequencing method based on real-time pyrophosphate[J]. Science, 1998, 281(5375): 363-365.
    Busato F, Dejeux E, El Abdalaoui H, et al. Quantitative DNA Methylation Analysis at Single-Nucleotide Resolution by Pyrosequencing[J]. Methods Mol Biol, 2018, 1708: 427-445.
    Kunze S. Quantitative Region-Specific DNA Methylation Analysis by the EpiTYPER™ Technology[J]. Methods Mol Biol, 2018, 1708: 515-535.
    Suchiman HE, Slieker RC, Kremer D, et al. Design, measurement and processing of region-specific DNA methylation assays: the mass spectrometry-based method EpiTYPER[J]. Frontiers in genetics, 2015, 6: 287-294.
    Ehrich M, Nelson MR, Stanssens P, et al. Quantitative high-throughput analysis of DNA methylation patterns by base-specific cleavage and mass spectrometry[J]. Proc Natl Acad Sci, 2005, 102(44): 15785-15790. doi: 10.1073/pnas.0507816102
    Coolen MW, Statham AL, Gardiner-Garden M, et al. Genomic profiling of CpG methylation and allelic specificity using quantitative high-throughput mass spectrometry: critical evaluation and improvements[J]. Nucleic acids research, 2007, 35(18): 119-125. doi: 10.1093/nar/gkm662
    Ehrich M, Turner J, Gibbs P, et al. Cytosine methylation profiling of cancer cell lines[J]. Proc Natl Acad Sci, 2008, 105(12): 4844-4849. doi: 10.1073/pnas.0712251105
    Docherty SJ, Davis OS, Haworth CM, et al. Bisulfite-based epityping on pooled genomic DNA provides an accurate estimate of average group DNA methylation[J]. Epigenetics & chromatin, 2009, 2(1): 3-10.
    Hussmann D, Hansen LL. Methylation-Sensitive High Resolution Melting (MS-HRM)[J]. Methods Mol Biol, 2018, 1708: 551-571.
    Frommer M, McDonald LE, Millar, et al. A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands[J]. Proc Natl Acad Sci, 1992, 89(5): 1827-1831. doi: 10.1073/pnas.89.5.1827
    Hsu HK, Weng YI, Hsu PY, et al. Detection of DNA methylation by MeDIP and MBDCap assays: an overview of techniques[J]. Methods Mol Biol, 2014, 1105: 61-70.
    Weber M, Hellmann I, Stadler MB, et al. Distribution, silencing potential and evolutionary impact of promoter DNA methylation in the human genome[J]. Nature genetics, 2007, 39(4): 457-466. doi: 10.1038/ng1990
    Weber M, Davies JJ, Wittig D, et al. Chromosome-wide and promoter-specific analyses identify sites of differential DNA methylation in normal and transformed human cells[J]. Nature genetics, 2005, 37(8): 853-862. doi: 10.1038/ng1598
    Down TA, Rakyan VK, Turner DJ, et al. A Bayesian deconvolution strategy for immunoprecipitation-based DNA methylome analysis[J]. Nature biotechnology, 2008, 26(7): 779-785. doi: 10.1038/nbt1414
    Keshet I, Schlesinger Y, Farkash S, et al. Evidence for an instructive mechanism of de novo methylation in cancer cells[J]. Nature genetics, 2006, 38(2): 149-153. doi: 10.1038/ng1719
    Zhang X, Yazaki J, Sundaresan A, et al. Genome-wide high-resolution mapping and functional analysis of DNA methylation in arabidopsis[J]. Cell, 2006, 126(6): 1189-1201. doi: 10.1016/j.cell.2006.08.003
    Zilberman D, Gehring M, Tran RK, et al. Genome-wide analysis of Arabidopsis thaliana DNA methylation uncovers an interdependence between methylation and transcription[J]. Nature genetics, 2007, 39(1): 61-69. doi: 10.1038/ng1929
    Zhao MT, Whyte JJ, Hopkins GM, et al. Methylated DNA immunoprecipitation and high-throughput sequencing (MeDIP-seq) using low amounts of genomic DNA[J]. Cellular reprogramming, 2014, 16(3): 175-184. doi: 10.1089/cell.2014.0002
    Serre D, Lee BH, Ting AH. MBD-isolated Genome Sequencing provides a high-throughput and comprehensive survey of DNA methylation in the human genome[J]. Nucleic acids research, 2010, 38(2): 391-399. doi: 10.1093/nar/gkp992
    Robinson MD, Stirzaker C, Statham AL, et al. Evaluation of affinity-based genome-wide DNA methylation data: effects of CpG density, amplification bias, and copy number variation[J]. Genome research, 2010, 20(12): 1719-1729. doi: 10.1101/gr.110601.110
    Weisenberger DJ, Trinh BN, Campan M, et al. DNA methylation analysis by digital bisulfite genomic sequencing and digital MethyLight[J]. Nucleic acids research, 2008, 36(14): 4689-4698. doi: 10.1093/nar/gkn455
    Li M, Chen WD, Papadopoulos N, et al. Sensitive digital quantification of DNA methylation in clinical samples[J]. Nature biotechnology, 2009, 27(9): 858-863. doi: 10.1038/nbt.1559
    Hao X, Luo H, Krawczyk M, et al. DNA methylation markers for diagnosis and prognosis of common cancers[J]. Proc Natl Acad Sci, 2017, 114(28): 7414-7419. doi: 10.1073/pnas.1703577114
    Van W L, Janssens L, Meeuws H, et al. Droplet digital PCR is an accurate method to assess methylation status on FFPE samples[J]. Epigenetics, 2018, 13(3): 207-213. doi: 10.1080/15592294.2018.1448679
    Barault L, Amatu A, Bleeker FE, et al. Digital PCR quantification of MGMT methylation refines prediction of clinical benefit from alkylating agents in glioblastoma and metastatic colorectal cancer[J]. Annals of oncology:official journal of the European Society for Medical Oncology, 2015, 26(9): 1994-1999. doi: 10.1093/annonc/mdv272
    Cui X, Cao L, Huang Y, et al. In vitro diagnosis of DNA methylation biomarkers with digital PCR in breast tumors[J]. The Analyst, 2018, 143(13): 3011-3020. doi: 10.1039/C8AN00205C
    Wu Z, Bai Y, Cheng Z, et al. Absolute quantification of DNA methylation using microfluidic chip-based digital PCR[J]. Biosensors & bioelectronics, 2017, 96: 339-344.
    Ladd AC, Aryee MJ, Ordway JM, et al. Comprehensive high-throughput arrays for relative methylation (CHARM)[J]. Current protocols in human genetics, 2010, 20: 1-19.
    Warnecke PM, Stirzaker C, Melki JR, et al. Detection and measurement of PCR bias in quantitative methylation analysis of bisulphite-treated DNA[J]. Nucleic acids research, 1997, 25(21): 4422-4426. doi: 10.1093/nar/25.21.4422
    Warnecke PM, Stirzaker C, Song J, et al. Identification and resolution of artifacts in bisulfite sequencing[J]. Methods, 2002, 27(2): 101-107. doi: 10.1016/S1046-2023(02)00060-9
    Hernández HG, Tse MY, Pang SC, et al. Optimizing methodologies for PCR-based DNA methylation analysis[J]. Biotechniques, 2013, 55(4): 181-197. doi: 10.2144/000114087
    Sant KE, Nahar MS, Dolinoy DC. DNA methylation screening and analysis[J]. Methods Mol Biol, 2012, 889: 385-406.
    Wang RY, Gehrke CW, Ehrlich M. Comparison of bisulfite modification of 5-methyldeoxycytidine and deoxycytidine residues[J]. Nucleic acids research, 1980, 8(20): 4777-4790. doi: 10.1093/nar/8.20.4777
    Pajares MJ, Palanca BC, Urtasun R, et al. Methods for analysis of specific DNA methylation status[J]. Methods, 2021, 187: 3-12. doi: 10.1016/j.ymeth.2020.06.021
    陈桂芳, 欧阳艳艳, 杨佳怡, 等. 核酸标准物质测量方法研究进展[J]. 计量科学与技术, 2021, 65(6): 25-33. doi: 10.12338/j.issn.2096-9015.2020.9022
    Meissner A, Mikkelsen TS, Gu H, et al. Genome-scale DNA methylation maps of pluripotent and differentiated cells[J]. Nature, 2008, 454(7205): 766-770. doi: 10.1038/nature07107
    Lister R, Pelizzola M, Dowen RH, et al. Human DNA methylomes at base resolution show widespread epigenomic differences[J]. Nature, 2009, 462(7271): 315-322. doi: 10.1038/nature08514
    高运华, 陈鸿飞, 盛灵慧, 等. DNA甲基化定量测量国际比对CCQM P94.2[J]. 中国测试, 2015(6): 47-51. doi: 10.11857/j.issn.1674-5124.2015.06.011
    牛春艳, 张永卓, 杨佳怡, 等. 基于数字PCR的质粒核酸标准物质合作定值研究[J]. 计量学报, 2021, 42(11): 1522-1527. doi: 10.3969/j.issn.1000-1158.2021.11.18
    Yang I, Kim SK, Burke DG, et al. An international comparability study on quantification of total methyl cytosine content[J]. Analytical biochemistry, 2009, 384(2): 288-295. doi: 10.1016/j.ab.2008.09.036
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