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© Research
Publication : Genome research

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

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in Genome research - 19 Jan 2021

Sun Z, Vaisvila R, Hussong LM, Yan B, Baum C, Saleh L, Samaranayake M, Guan S, Dai N, Corrêa IR, Pradhan S, Davis TB, Evans TC, Ettwiller LM,

Link to Pubmed [PMID] – 33468551

Link to DOI – 10.1101/gr.265306.120

Genome Res 2021 Jan; ():

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.

https://pubmed.ncbi.nlm.nih.gov/33468551