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Non-destructive enzymatic deamination enables single molecule long read sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single base resolution

View ORCID ProfileZhiyi Sun, Romualdas Vaisvila, View ORCID ProfileBo Yan, Chloe Baum, View ORCID ProfileLana Saleh, Mala Samaranayake, Shengxi Guan, Nan Dai, Ivan R. Corrêa Jr., Sriharsa Pradhan, Theodore B. Davis, Thomas C. Evans Jr., View ORCID ProfileLaurence M. Ettwiller
doi: https://doi.org/10.1101/2019.12.20.885061
Zhiyi Sun
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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  • For correspondence: sunz@neb.com ettwiller@neb.com
Romualdas Vaisvila
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Bo Yan
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Chloe Baum
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Lana Saleh
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Mala Samaranayake
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Shengxi Guan
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Nan Dai
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Ivan R. Corrêa Jr.
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Sriharsa Pradhan
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Theodore B. Davis
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Thomas C. Evans Jr.
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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Laurence M. Ettwiller
1New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938 USA
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  • ORCID record for Laurence M. Ettwiller
  • For correspondence: sunz@neb.com ettwiller@neb.com
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Abstract

The predominant methodology for DNA methylation analysis relies on the chemical deamination by sodium bisulfite of unmodified cytosine to uracil to permit the differential readout of methylated cytosines. Bisulfite treatment damages the DNA leading to fragmentation and loss of long-range methylation information. To overcome this limitation of bisulfite treated DNA we applied a new enzymatic deamination approach, termed EM-seq (Enzymatic Methyl-seq) to long-range sequencing technologies. Our methodology, named LR-EM-seq (Long Range Enzymatic Methyl-seq) preserves the integrity of DNA allowing long-range methylation profiling of 5-mC and 5-hmC over several kilobases of genomic DNA. When applied to known differentially methylated regions (DMR), LR-EM-seq achieves phasing of over 5 kb resulting in broader and better defined DMRs compared to previously reported. This result demonstrated the importance of phasing methylation for biologically relevant questions and the applicability of LR-EM-seq for long range epigenetic analysis at single molecule and single nucleotide resolution.

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Posted December 23, 2019.
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Non-destructive enzymatic deamination enables single molecule long read sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single base resolution
Zhiyi Sun, Romualdas Vaisvila, Bo Yan, Chloe Baum, Lana Saleh, Mala Samaranayake, Shengxi Guan, Nan Dai, Ivan R. Corrêa Jr., Sriharsa Pradhan, Theodore B. Davis, Thomas C. Evans Jr., Laurence M. Ettwiller
bioRxiv 2019.12.20.885061; doi: https://doi.org/10.1101/2019.12.20.885061
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Non-destructive enzymatic deamination enables single molecule long read sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single base resolution
Zhiyi Sun, Romualdas Vaisvila, Bo Yan, Chloe Baum, Lana Saleh, Mala Samaranayake, Shengxi Guan, Nan Dai, Ivan R. Corrêa Jr., Sriharsa Pradhan, Theodore B. Davis, Thomas C. Evans Jr., Laurence M. Ettwiller
bioRxiv 2019.12.20.885061; doi: https://doi.org/10.1101/2019.12.20.885061

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