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

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 methylatio...

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Autores principales: Sun, Zhiyi, Vaisvila, Romualdas, Hussong, Laura-Madison, Yan, Bo, Baum, Chloé, Saleh, Lana, Samaranayake, Mala, Guan, Shengxi, Dai, Nan, Corrêa, Ivan R., Pradhan, Sriharsa, Davis, Theodore B., Evans, Thomas C., Ettwiller, Laurence M.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Cold Spring Harbor Laboratory Press 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7849414/
https://www.ncbi.nlm.nih.gov/pubmed/33468551
http://dx.doi.org/10.1101/gr.265306.120
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author Sun, Zhiyi
Vaisvila, Romualdas
Hussong, Laura-Madison
Yan, Bo
Baum, Chloé
Saleh, Lana
Samaranayake, Mala
Guan, Shengxi
Dai, Nan
Corrêa, Ivan R.
Pradhan, Sriharsa
Davis, Theodore B.
Evans, Thomas C.
Ettwiller, Laurence M.
author_facet Sun, Zhiyi
Vaisvila, Romualdas
Hussong, Laura-Madison
Yan, Bo
Baum, Chloé
Saleh, Lana
Samaranayake, Mala
Guan, Shengxi
Dai, Nan
Corrêa, Ivan R.
Pradhan, Sriharsa
Davis, Theodore B.
Evans, Thomas C.
Ettwiller, Laurence M.
author_sort Sun, Zhiyi
collection PubMed
description 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 enzymatic methyl-seq (EM-seq), to long-range sequencing technologies. Our methodology, named long-read enzymatic modification sequencing (LR-EM-seq), preserves the integrity of DNA, allowing long-range methylation profiling of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) over multikilobase length of genomic DNA. When applied to known differentially methylated regions (DMRs), LR-EM-seq achieves phasing of >5 kb, resulting in broader and better defined DMRs compared with that previously reported. This result showed 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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spelling pubmed-78494142021-02-04 Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution Sun, Zhiyi Vaisvila, Romualdas Hussong, Laura-Madison Yan, Bo Baum, Chloé Saleh, Lana Samaranayake, Mala Guan, Shengxi Dai, Nan Corrêa, Ivan R. Pradhan, Sriharsa Davis, Theodore B. Evans, Thomas C. Ettwiller, Laurence M. Genome Res Method 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 enzymatic methyl-seq (EM-seq), to long-range sequencing technologies. Our methodology, named long-read enzymatic modification sequencing (LR-EM-seq), preserves the integrity of DNA, allowing long-range methylation profiling of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) over multikilobase length of genomic DNA. When applied to known differentially methylated regions (DMRs), LR-EM-seq achieves phasing of >5 kb, resulting in broader and better defined DMRs compared with that previously reported. This result showed 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. Cold Spring Harbor Laboratory Press 2021-02 /pmc/articles/PMC7849414/ /pubmed/33468551 http://dx.doi.org/10.1101/gr.265306.120 Text en © 2021 Sun et al.; Published by Cold Spring Harbor Laboratory Press http://creativecommons.org/licenses/by-nc/4.0/ This article, published in Genome Research, is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.
spellingShingle Method
Sun, Zhiyi
Vaisvila, Romualdas
Hussong, Laura-Madison
Yan, Bo
Baum, Chloé
Saleh, Lana
Samaranayake, Mala
Guan, Shengxi
Dai, Nan
Corrêa, Ivan R.
Pradhan, Sriharsa
Davis, Theodore B.
Evans, Thomas C.
Ettwiller, Laurence M.
Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution
title Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution
title_full Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution
title_fullStr Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution
title_full_unstemmed Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution
title_short Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution
title_sort nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution
topic Method
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7849414/
https://www.ncbi.nlm.nih.gov/pubmed/33468551
http://dx.doi.org/10.1101/gr.265306.120
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