Highlights
- •Evaluation of methylation age markers using microarray data and targeted sequencing revealed a set of 11 ‘optimal’ markers.
- •The prediction model showed high prediction accuracy in both a UK (MAE = 3.3 years) and Spanish sample cohort (MAE = 3.8 years).
- •Prediction accuracy improved for under 55-year-olds (MAE = 2.6), with 81% predicting with an error of less than 4 years.
- •The accuracy of DNA methylation quantification and age prediction was retained down to 5 ng of DNA input (~ 1 ng in PCR stage).
Abstract
The estimation of chronological age from biological fluids has been an important quest
for forensic scientists worldwide, with recent approaches exploiting the variability
of DNA methylation patterns with age in order to develop the next generation of forensic
‘DNA intelligence’ tools for this application. Drawing from the conclusions of previous
work utilising massively parallel sequencing (MPS) for this analysis, this work introduces
a DNA methylation-based age estimation method for blood that exhibits the best combination
of prediction accuracy and sensitivity reported to date. Statistical evaluation of
markers from 51 studies using microarray data from over 4000 individuals, followed
by validation using in-house generated MPS data, revealed a final set of 11 markers
with the greatest potential for accurate age estimation from minimal DNA material.
Utilising an algorithm based on support vector machines, the proposed model achieved
an average error (MAE) of 3.3 years, with this level of accuracy retained down to
5 ng of starting DNA input (~ 1 ng PCR input). The accuracy of the model was retained
(MAE = 3.8 years) in a separate test set of 88 samples of Spanish origin, while predictions
for donors of greater forensic interest (< 55 years of age) displayed even higher
accuracy (MAE = 2.6 years). Finally, no sex-related bias was observed for this model,
while there were also no signs of variation observed between control and disease-associated
populations for schizophrenia, rheumatoid arthritis, frontal temporal dementia and
progressive supranuclear palsy in microarray data relating to the 11 markers.
Keywords
To read this article in full you will need to make a payment
Purchase one-time access:
Academic & Personal: 24 hour online accessCorporate R&D Professionals: 24 hour online accessOne-time access price info
- For academic or personal research use, select 'Academic and Personal'
- For corporate R&D use, select 'Corporate R&D Professionals'
Subscribe:
Subscribe to Forensic Science International: GeneticsAlready a print subscriber? Claim online access
Already an online subscriber? Sign in
Register: Create an account
Institutional Access: Sign in to ScienceDirect
References
- Isolation and identification of age-related DNA methylation markers for forensic age-prediction.Forensic Sci. Int. Genet. 2014; 11: 117-125
- A comparison of feature selection and classification methods in DNA methylation studies using the Illumina Infinium platform.BMC Bioinform. 2012; 13: 59
- DNA methylation age of human tissues and cell types.Genome Biol. 2013; 14: 115
- Principles and challenges of genomewide DNA methylation analysis.Nat. Rev. Genet. 2010; 11: 191-203
- A comprehensive overview of Infinium HumanMethylation450 data processing.Brief. Bioinform. 2014; 15: 929-941
- Epigenetic-aging-signature to determine age in different tissues.Aging. 2011; 3: 1018-1027
- Focused, high accuracy 5-methylcytosine quantitation with base resolution by benchtop next-generation sequencing.Epigenet. Chromatin. 2013; 6: 33
- Examination of DNA methylation status of the ELOVL2 marker may be useful for human age prediction in forensic science.Forensic Sci. Int. Genet. 2015; 14: 161-167
- DNA methylation and cancer.J. Clin. Oncol. 2004; 22: 4632-4642
- Epigenetic age of the pre-frontal cortex is associated with neuritic plaques, amyloid load, and Alzheimer’s disease related cognitive functioning.Aging. 2015; 7 (Albany NY): 1198-1211
- Epigenetic studies in Alzheimer’s disease: current findings, caveats, and considerations for future studies.Am. J. Med. Genet. B Neuropsychiatr. Genet. 2013; 162B: 789-799
- A cross-brain regions study of ANK1 DNA methylation in different neurodegenerative diseases.Neurobiol. Aging. 2019; 74: 70-76
- Huntington’s disease accelerates epigenetic aging of human brain and disrupts DNA methylation levels.Aging. 2016; 8 (Albany NY): 1485-1512
- Increased epigenetic age and granulocyte counts in the blood of Parkinson’s disease patients.Aging. 2015; 7 (Albany NY): 1130-1142
- Epigenetic clock for skin and blood cells applied to Hutchinson Gilford Progeria Syndrome and ex vivo studies.Aging. 2018; 10 (Albany NY): 1758-1775
- Accelerated epigenetic aging in Werner syndrome.Aging. 2017; 9: 1143-1152
- Tobacco-smoking-related differential DNA methylation: 27K discovery and replication.Am. J. Hum. Genet. 2011; 88: 450-457
- Cigarette smoking significantly alters sperm DNA methylation patterns.Andrology. 2017; 5: 1089-1099
- Cigarette smoking and DNA methylation.Front. Genet. 2013; 4: 132
- Differential DNA methylation in relation to age and health risks of obesity.Int. J. Mol. Sci. 2015; 16: 16816-16832
- Genome-wide analysis reveals DNA methylation markers that vary with both age and obesity.Gene. 2014; 548: 61-67
- An epigenetic biomarker of aging for lifespan and healthspan.Aging. 2018; 10 (Albany NY): 573-591
- Socioeconomic position and DNA methylation age acceleration across the life course.Am. J. Epidemiol. 2018; 187: 2346-2354
- Genome-wide analysis of DNA methylation in relation to socioeconomic status during development and early adulthood.Am. J. Phys. Anthropol. 2019; 169: 3-11
- Biomarkers of aging.Exp. Gerontol. 1988; 23: 1-3
- DNA methylation GrimAge strongly predicts lifespan and healthspan.Aging. 2019; 11 (Albany NY): 303-327
- DNA methylation-based forensic age prediction using artificial neural networks and next generation sequencing.Forensic Sci. Int. Genet. 2017; 28: 225-236
- DNA methylation-based age prediction using massively parallel sequencing data and multiple machine learning models.Forensic Sci. Int. Genet. 2018; 37: 215-226
- The relationship of DNA methylation with age, gender and genotype in twins and healthy controls.PLoS One. 2009; 4e6767
- Human aging-associated DNA hypermethylation occurs preferentially at bivalent chromatin domains.Genome Res. 2010; 20: 434-439
- Age-dependent DNA methylation of genes that are suppressed in stem cells is a hallmark of cancer.Genome Res. 2010; 20: 440-446
- Epigenetic predictor of age.PLoS One. 2011; 6e14821
- Specific age-associated DNA methylation changes in human dermal fibroblasts.PLoS One. 2011; 6e16679
- Distinct DNA methylation changes highly correlated with chronological age in the human brain.Hum. Mol. Genet. 2011; 20: 1164-1172
- Evidence for age-related and individual-specific changes in DNA methylation profile of mononuclear cells during early immune development in humans.Epigenetics. 2011; 6: 1085-1094
- Epigenome-wide scans identify differentially methylated regions for age and age-related phenotypes in a healthy ageing population.PLoS Genet. 2012; 8e1002629
- Aging effects on DNA methylation modules in human brain and blood tissue.Genome Biol. 2012; 13: R97
- Methylation of ELOVL2 gene as a new epigenetic marker of age.Aging Cell. 2012; 11: 1132-1134
- Age-associated DNA methylation in pediatric populations.Genome Res. 2012; 22: 623-632
- DNA methylation signatures in development and aging of the human prefrontal cortex.Am. J. Hum. Genet. 2012; 90: 260-272
- Age-associated epigenetic drift: implications, and a case of epigenetic thrift?.Hum. Mol. Genet. 2013; 22: R7-R15
- Differential DNA methylation with age displays both common and dynamic features across human tissues that are influenced by CpG landscape.Genome Biol. 2013; 14: R102
- Genome-wide methylation profiles reveal quantitative views of human aging rates.Mol. Cell. 2013; 49: 359-367
- DNA methylome profiling using neonatal dried blood spot samples: a proof-of-principle study.Mol. Genet. Metab. 2013; 108: 225-231
- Continuous aging of the human DNA methylome throughout the human lifespan.PLoS One. 2013; 8e67378
- Genome-wide DNA methylation changes with age in disease-free human skeletal muscle.Aging Cell. 2014; 13: 360-366
- Longitudinal, genome-scale analysis of DNA methylation in twins from birth to 18 months of age reveals rapid epigenetic change in early life and pair-specific effects of discordance.Genome Biol. 2013; 14: R42
- Cross-sectional and longitudinal changes in DNA methylation with age: an epigenome-wide analysis revealing over 60 novel age-associated CpG sites.Hum. Mol. Genet. 2014; 23: 1186-1201
- Aging of blood can be tracked by DNA methylation changes at just three CpG sites.Genome Biol. 2014; 15: R24
- Genome-wide age-related changes in DNA methylation and gene expression in human PBMCs.Age. 2014; 36: 9648
- Ageing-associated changes in the human DNA methylome: genomic locations and effects on gene expression.BMC Genom. 2015; 16: 179
- A methylome-wide study of aging using massively parallel sequencing of the methyl-CpG-enriched genomic fraction from blood in over 700 subjects.Hum. Mol. Genet. 2014; 23: 1175-1185
- Improved age determination of blood and teeth samples using a selected set of DNA methylation markers.Epigenetics. 2015; 10: 922-930
- Developing a DNA methylation assay for human age prediction in blood and bloodstain.Forensic Sci. Int. Genet. 2015; 17: 129-136
- Epigenetic age signatures in the forensically relevant body fluid of semen: a preliminary study.Forensic Sci. Int. Genet. 2015; 19: 28-34
- Evaluation of DNA methylation markers and their potential to predict human aging.Electrophoresis. 2015; 36: 1775-1780
- Age-related DNA methylation changes for forensic age-prediction.Int. J. Leg. Med. 2015; 129: 237-244
- Association of DNA methylation with age, gender, and smoking in an Arab population.Clin. Epigenet. 2015; 7: 6
- A novel strategy for forensic age prediction by DNA methylation and support vector regression model.Sci. Rep. 2015; 5: 17788
- Age-associated DNA methylation changes in immune genes, histone modifiers and chromatin remodeling factors within 5 years after birth in human blood leukocytes.Clin. Epigenet. 2015; 7: 34
- The transcriptional landscape of age in human peripheral blood.Nat. Commun. 2015; 6: 8570
- Human age estimation from blood using mRNA, DNA methylation, DNA rearrangement, and telomere length.Forensic Sci. Int. Genet. 2016; 24: 33-43
- Identification and evaluation of age-correlated DNA methylation markers for forensic use.Forensic Sci. Int. Genet. 2016; 23: 64-70
- Development of a methylation marker set for forensic age estimation using analysis of public methylation data and the Agena Bioscience EpiTYPER system.Forensic Sci. Int. Genet. 2016; 24: 65-74
- Cytomegalovirus infection accelerates epigenetic aging.Exp. Gerontol. 2015; 72: 227-229
- Simplified assay for epigenetic age estimation in whole blood of adults.Front. Genet. 2016; 7: 126
- An epigenetic clock for gestational age at birth based on blood methylation data.Genome Biol. 2016; 17: 206
- Epigenetic drift in the aging genome: a ten-year follow-up in an elderly twin cohort.Int. J. Epidemiol. 2016; 45: 1146-1158
- DNA methylation-based age prediction from saliva: high age predictability by combination of 7 CpG markers.Forensic Sci. Int. Genet. 2017; 29: 118-125
- Accelerated placental aging in early onset preeclampsia pregnancies identified by DNA methylation.Epigenomics. 2017; 9: 279-289
- Independent validation of DNA-based approaches for age prediction in blood.Forensic Sci. Int. Genet. 2017; 29: 250-256
- Methylome-wide association study of whole blood DNA in the Norfolk Island isolate identifies robust loci associated with age.Aging. 2017; 9 (Albany, NY): 753-768
- The emerging landscape of dynamic DNA methylation in early childhood.BMC Genom. 2017; 18: 25
- NCBI GEO: archive for functional genomics data sets--update.Nucleic Acids Res. 2013; 41 (Database issue): D991-D995
- A BRCA1-mutation associated DNA methylation signature in blood cells predicts sporadic breast cancer incidence and survival.Genome Med. 2014; 6: 47
- Sequence overlap between autosomal and sex-linked probes on the Illumina HumanMethylation27 microarray.Genomics. 2011; 97: 214-222
- HIV-1 infection accelerates age according to the epigenetic clock.J. Infect. Dis. 2015; 212: 1563-1573
- Epigenome-wide association data implicate DNA methylation as an intermediary of genetic risk in rheumatoid arthritis.Nat. Biotechnol. 2013; 31: 142-147
- Genome wide peripheral blood leukocyte DNA methylation microarrays identified a single association with inflammatory bowel diseases.Inflamm. Bowel Dis. 2012; 18 (p. 10.1002/ibd.22956)
- Differential methylation of the TRPA1 promoter in pain sensitivity.Nat. Commun. 2014; 5: 2978
- An epigenetic clock analysis of race/ethnicity, sex, and coronary heart disease.Genome Biol. 2016; 17: 171
- Comparison of beta-value and M-value methods for quantifying methylation levels by microarray analysis.BMC Bioinform. 2010; 11: 587
- Quantifiler™ HP and Trio DNA Quantification Kits User Guide.Thermo Fisher Scientific, 2017
- MethylEdgeTM Bisulfite Conversion System Instructions for use of Product N1301.Promega Corporation, 2013
- Bisulfite conversion of DNA: performance comparison of different kits and methylation quantitation of epigenetic biomarkers that have the potential to be used in non-invasive prenatal testing.PLOS ONE. 2015; 10e0135058
- MethPrimer: designing primers for methylation PCRs.Bioinformatics. 2002; 18: 1427-1431
- Ensembl 2020.Nucleic Acids Res. 2020; 48: D682-D688
- Chronological age prediction based on DNA methylation: massive parallel sequencing and random forest regression.Forensic Sci. Int. Genet. 2017; 31: 19-28
- MinElute® PCR Purification Kit Quick Start Protocol.Qiagen Sample and Assay Technologies, 2011
L. Technologies, User Guide: Qubit® dsDNA HS Assay Kits for use with the Qubit® Fluorometer (all models). Life Technologies Molecular Probes, 2015.
N.E. BioLabs, NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® (E7645, E7103) Instruction Manual, N.E. BioLabs, Editor.
- KAPA Hyper Prep Kit Technical Data Sheet KR0961 – v5.16.KAPA Biosystems, 2016
- KAPA Library Quantification Kit Technical Data Sheet KR0405 – v8.17.KAPA Biosystems, 2017
- Fast and accurate short read alignment with Burrows–Wheeler transform.Bioinformatics. 2009; 25: 1754-1760
- The sequence alignment/map format and SAMtools.Bioinformatics. 2009; 25: 2078-2079
- The genome analysis toolkit: a mapreduce framework for analyzing next-generation DNA sequencing data.Genome Res. 2010; 20: 1297-1303
- DNA methylation in mammals.Cold Spring Harb. Perspect. Biol. 2014; 6: a019133
- Methylation of CpG sequences in eukaryotic DNA.FEBS Lett. 1981; 124: 67-71
R.C. Team, R: A Language and Environment for Statistical Computing. R Foundation for Statistical Computing, Vienna, Austria, 2020.
- Building predictive models in R using the caret package.J. Stat. Softw. 2008; 28: 26
- EWAS data hub: a resource of DNA methylation array data and metadata.Nucleic Acids Res. 2020; 48: D890-D895
- PANTHER version 14: more genomes, a new PANTHER GO-slim and improvements in enrichment analysis tools.Nucleic Acids Res. 2019; 47: D419-D426
- Protocol update for large-scale genome and gene function analysis with the PANTHER classification system (v.14.0).Nat. Protoc. 2019; 14: 703-721
- PANTHER pathway: an ontology-based pathway database coupled with data analysis tools.Methods Mol. Biol. 2009; 563: 123-140
- Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists.Nucleic Acids Res. 2009; 37: 1-13
- Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources.Nat. Protoc. 2009; 4: 44-57
- Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists.Nucleic Acids Res. 2009; 37: 1-13
- New approach for understanding genome variations in KEGG.Nucleic Acids Res. 2019; 47: D590-D595
- The genetic association database.Nat. Genet. 2004; 36: 431-432
- A comparison of forensic age prediction models using data from four DNA methylation technologies.Front. Genet. 2020; 11: 932
- Forensic age prediction for dead or living samples by use of methylation-sensitive high resolution melting.Leg. Med. 2016; 21: 5-10
- Proof of concept study of age-dependent DNA methylation markers across different tissues by massive parallel sequencing.Forensic Sci. Int. Genet. 2018; 36: 152-159
H. Office, National DNA Database Strategy Board Biennial Report 2018–2020, 2020.
- Female marrow donors increase the risk of acute graft-versus-host disease: effect of donor age and parity and analysis of cell subpopulations in the donor marrow inoculum.Br. J. Haematol. 1986; 63: 231-239
- DNA methylation signatures in peripheral blood strongly predict all-cause mortality.Nat. Commun. 2017; 8: 14617
- Obesity accelerates epigenetic aging in middle-aged but not in elderly individuals.Clin. Epigenet. 2017; 9: 20
- The hallmarks of aging.Cell. 2013; 153: 1194-1217
- Metabolic control of longevity.Cell. 2016; 166: 802-821
- Organization of the mammalian metabolome according to organ function, lineage specialization, and longevity.Cell Metab. 2015; 22: 332-343
- Promoting health and longevity through diet: from model organisms to humans.Cell. 2015; 161: 106-118
- Development of a forensically useful age prediction method based on DNA methylation analysis.Forensic Sci. Int. Genet. 2015; 17: 173-179
Article info
Publication history
Published online: November 23, 2021
Accepted:
November 17,
2021
Received in revised form:
October 19,
2021
Received:
June 2,
2021
Identification
Copyright
© 2021 Elsevier B.V. All rights reserved.
