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Pros and cons of methylation-based enrichment methods for ancient DNA.

Seguin-Orlando A, Gamba C, Der Sarkissian C, Ermini L, Louvel G, Boulygina E, Sokolov A, Nedoluzhko A, Lorenzen ED, Lopez P, McDonald HG, Scott E, Tikhonov A, Stafford TW, Alfarhan AH, Alquraishi SA, Al-Rasheid KA, Shapiro B, Willerslev E, Prokhortchouk E, Orlando L - Sci Rep (2015)

Bottom Line: The recent discovery that DNA methylation survives in fossil material provides an opportunity for novel molecular approaches in palaeogenomics.MBD enrichment, however, appears principally biased towards the recovery of CpG-rich and long DNA templates and is limited by the fast post-mortem cytosine deamination rates of methylated epialleles.This work represents an essential step toward the characterization of ancient methylation signatures, which will help understanding the role of epigenetic changes in past environmental and cultural transitions.

View Article: PubMed Central - PubMed

Affiliation: 1] Centre for GeoGenetics, Natural History Museum of Denmark, Øster Voldgade 5-7, 1350K Copenhagen, Denmark [2] National High-throughput DNA Sequencing Centre, Øster Farimagsgade 2D, 1353K Copenhagen, Denmark.

ABSTRACT
The recent discovery that DNA methylation survives in fossil material provides an opportunity for novel molecular approaches in palaeogenomics. Here, we apply to ancient DNA extracts the probe-independent Methylated Binding Domains (MBD)-based enrichment method, which targets DNA molecules containing methylated CpGs. Using remains of a Palaeo-Eskimo Saqqaq individual, woolly mammoths, polar bears and two equine species, we confirm that DNA methylation survives in a variety of tissues, environmental contexts and over a large temporal range (4,000 to over 45,000 years before present). MBD enrichment, however, appears principally biased towards the recovery of CpG-rich and long DNA templates and is limited by the fast post-mortem cytosine deamination rates of methylated epialleles. This method, thus, appears only appropriate for the analysis of ancient methylomes from very well preserved samples, where both DNA fragmentation and deamination have been limited. This work represents an essential step toward the characterization of ancient methylation signatures, which will help understanding the role of epigenetic changes in past environmental and cultural transitions.

No MeSH data available.


Related in: MedlinePlus

Molecular characteristics of DNA library inserts and post-mortem DNA degradation.(A) Percentage of CpGs in the nuclear DNA, normalized by read length. (B) Length of DNA library inserts. (C) Cytosine deamination probability in double-stranded DNA context (δd). (D) Cytosine deamination probability in single-stranded DNA contexts (δs). DNA degradation parameters were calculated using mapDamage 227. A minimum threshold of 2,000 reads was applied to all data shown (mapping quality filtered, MQ25) so as to retrieve unbiased distributions. MBD+ = MBD enriched fraction. MBD− = uncaptured fraction.
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f3: Molecular characteristics of DNA library inserts and post-mortem DNA degradation.(A) Percentage of CpGs in the nuclear DNA, normalized by read length. (B) Length of DNA library inserts. (C) Cytosine deamination probability in double-stranded DNA context (δd). (D) Cytosine deamination probability in single-stranded DNA contexts (δs). DNA degradation parameters were calculated using mapDamage 227. A minimum threshold of 2,000 reads was applied to all data shown (mapping quality filtered, MQ25) so as to retrieve unbiased distributions. MBD+ = MBD enriched fraction. MBD− = uncaptured fraction.

Mentions: We next evaluated whether endogenous DNA fragments recovered in MBD+ and MBD− fractions exhibited similar molecular properties. Only samples showing at least 2,000 high-quality hits mapping uniquely against their respective reference genome (MQ25 filter) were considered so as to enable reliable estimates of base compositional bias. We implemented linear mixed models analyses to assess statistical differences within the two fractions for %GC content, CpG densities and the size of library inserts. We found that MBD+ fractions were significantly enriched in endogenous reads carrying higher %GC contents and CpG densities, with the exception of a single sample (MZh_1, Figs 1C and 3A; Supplementary Fig. S1; Supplementary Tables S4.1 and S4.2).


Pros and cons of methylation-based enrichment methods for ancient DNA.

Seguin-Orlando A, Gamba C, Der Sarkissian C, Ermini L, Louvel G, Boulygina E, Sokolov A, Nedoluzhko A, Lorenzen ED, Lopez P, McDonald HG, Scott E, Tikhonov A, Stafford TW, Alfarhan AH, Alquraishi SA, Al-Rasheid KA, Shapiro B, Willerslev E, Prokhortchouk E, Orlando L - Sci Rep (2015)

Molecular characteristics of DNA library inserts and post-mortem DNA degradation.(A) Percentage of CpGs in the nuclear DNA, normalized by read length. (B) Length of DNA library inserts. (C) Cytosine deamination probability in double-stranded DNA context (δd). (D) Cytosine deamination probability in single-stranded DNA contexts (δs). DNA degradation parameters were calculated using mapDamage 227. A minimum threshold of 2,000 reads was applied to all data shown (mapping quality filtered, MQ25) so as to retrieve unbiased distributions. MBD+ = MBD enriched fraction. MBD− = uncaptured fraction.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4488743&req=5

f3: Molecular characteristics of DNA library inserts and post-mortem DNA degradation.(A) Percentage of CpGs in the nuclear DNA, normalized by read length. (B) Length of DNA library inserts. (C) Cytosine deamination probability in double-stranded DNA context (δd). (D) Cytosine deamination probability in single-stranded DNA contexts (δs). DNA degradation parameters were calculated using mapDamage 227. A minimum threshold of 2,000 reads was applied to all data shown (mapping quality filtered, MQ25) so as to retrieve unbiased distributions. MBD+ = MBD enriched fraction. MBD− = uncaptured fraction.
Mentions: We next evaluated whether endogenous DNA fragments recovered in MBD+ and MBD− fractions exhibited similar molecular properties. Only samples showing at least 2,000 high-quality hits mapping uniquely against their respective reference genome (MQ25 filter) were considered so as to enable reliable estimates of base compositional bias. We implemented linear mixed models analyses to assess statistical differences within the two fractions for %GC content, CpG densities and the size of library inserts. We found that MBD+ fractions were significantly enriched in endogenous reads carrying higher %GC contents and CpG densities, with the exception of a single sample (MZh_1, Figs 1C and 3A; Supplementary Fig. S1; Supplementary Tables S4.1 and S4.2).

Bottom Line: The recent discovery that DNA methylation survives in fossil material provides an opportunity for novel molecular approaches in palaeogenomics.MBD enrichment, however, appears principally biased towards the recovery of CpG-rich and long DNA templates and is limited by the fast post-mortem cytosine deamination rates of methylated epialleles.This work represents an essential step toward the characterization of ancient methylation signatures, which will help understanding the role of epigenetic changes in past environmental and cultural transitions.

View Article: PubMed Central - PubMed

Affiliation: 1] Centre for GeoGenetics, Natural History Museum of Denmark, Øster Voldgade 5-7, 1350K Copenhagen, Denmark [2] National High-throughput DNA Sequencing Centre, Øster Farimagsgade 2D, 1353K Copenhagen, Denmark.

ABSTRACT
The recent discovery that DNA methylation survives in fossil material provides an opportunity for novel molecular approaches in palaeogenomics. Here, we apply to ancient DNA extracts the probe-independent Methylated Binding Domains (MBD)-based enrichment method, which targets DNA molecules containing methylated CpGs. Using remains of a Palaeo-Eskimo Saqqaq individual, woolly mammoths, polar bears and two equine species, we confirm that DNA methylation survives in a variety of tissues, environmental contexts and over a large temporal range (4,000 to over 45,000 years before present). MBD enrichment, however, appears principally biased towards the recovery of CpG-rich and long DNA templates and is limited by the fast post-mortem cytosine deamination rates of methylated epialleles. This method, thus, appears only appropriate for the analysis of ancient methylomes from very well preserved samples, where both DNA fragmentation and deamination have been limited. This work represents an essential step toward the characterization of ancient methylation signatures, which will help understanding the role of epigenetic changes in past environmental and cultural transitions.

No MeSH data available.


Related in: MedlinePlus