NEBNext® Enzymatic Methyl-seq Kit
Product informationCode | Name | Size | Quantity | Price | |
---|---|---|---|---|---|
E7120S |
NEBNext Enzymatic Methyl-seq Kit (EM-seq) |
24 Assays | - | Unavailable in your region | |
E7120L |
NEBNext Enzymatic Methyl-seq Kit (EM-seq) |
96 Assays | - | Unavailable in your region |
NEBNext® Enzymatic Methyl-seq Kit
For enzymatic fragmentation of DNA for EM-seq, NEBNext UltraShear™ (NEB #M7634) is now available.
Product Introduction
NEBNext Enzymatic Methyl-seq (EM-seq™) is a new method for identification of 5-mC and 5-hmC.
While bisulfite sequencing has been the gold standard for methylome analysis, this conversion treatment damages DNA, resulting in fragmentation, loss and bias.
In contrast, the highly effective enzyme-based conversion in the NEBNext Enzymatic Methyl-seq Kit minimizes damage to DNA and, with the supplied NEBNext Ultra™ II library preparation workflow reagents, produces high quality libraries that enable superior detection of 5-mC and 5-hmC from fewer sequencing reads.
- Superior sensitivity of detection of 5-mC and 5-hmC
- Greater mapping efficiency
- More uniform GC coverage
- Detection of more CpGs with fewer sequence reads
- Uniform dinucleotide distribution
- High-efficiency library preparation, with larger library insert sizes
- Conversion module also available separately
Enzymatic fragmentation of DNA compatible with EM-seq workflows can be achieved using NEBNext UltraShear (NEB #M7634).
View or download extensive EM-seq performance data in our Technical Note.
For specific detection of 5hmC, the NEBNext Enzymatic E5hmC-seq Kit (NEB #E3350) is now also available.
Catalog # | Size | Concentration |
---|---|---|
E7120S | 24.0 reactions | |
E7120L | 96.0 reactions |
- Product Information
- Protocols, Manuals & Usage
- FAQs & Troubleshooting
- Citations & Technical Literature
- Quality, Safety & Legal
- Other Products You May Be Interested In
Product Information
Description
View or download extensive performance data in our Technical Note.
The NEBNext Enzymatic Methyl-seq Kit provides a high-performance enzyme-based alternative to bisulfite conversion for methylome analysis using Illumina® sequencing.
Libraries are prepared using as little as 10 ng input DNA and the supplied NEBNext Ultra II reagents and the optimized EM-seq Adaptor. TET2 then oxidizes 5-mC and 5-hmC, providing protection from deamination by APOBEC in the next step. In contrast, unmodified cytosines are deaminated to uracils. Libraries are then amplified using a NEBNext master mix formulation of Q5U® (a modified version of Q5® High-Fidelity DNA Polymerase), and sequenced using Illumina instrumentation.
The consistently high conversion performance and minimized DNA damage with the EM-seq protocol, in combination with highly efficient Ultra II library prep, result in superior detection of CpGs with fewer sequencing reads.
NEBNext UltraShear (NEB #M7634) has been optimized for enzymatic fragmentation of DNA compatible with EM-seq workflows, and protocols are available in the NEBNext UltraShear product manual.
Features:
- Superior sensitivity of detection of 5-mC and 5-hmC
- Greater mapping efficiency
- More uniform GC coverage
- Detection of more CpGs with fewer sequence reads
- Uniform dinucleotide distribution
- High-efficiency library preparation, with larger library insert sizes
- Conversion module also available separately
For specific detection of 5hmC, the NEBNext Enzymatic E5hmC-seq Kit (NEB #E3350) is now also available.
NEBNext UltraShear can be used for enzymatic DNA fragmentation in the EM-seq workflow.
- This product is related to the following categories:
- Automation for NEBNext® NGS Library Prep Products,
- Methylome Analysis Products,
- Epigenetic Analysis Products,
- Epigenetics Products,
- This product can be used in the following applications:
- DNA Methylation Analysis,
- Bisulfite Conversion,
- Epigenetics, Methylated DNA Analysis Products
Kit Components
Kit Components
The following reagents are supplied with this product:
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Properties & Usage
Materials Required but not Supplied
- Covaris® S2 instrument or other fragmentation equipment
- PCR strip tubes
- Formamide (Sigma #F9037-100 ml) or 0.1 N NaOH
- 80% Ethanol • 0.1X TE, pH 8.0
- Nuclease-free Water
- Magnetic rack/stand, such as NEBNext Magnetic Separation Rack (NEB #S1515)
- PCR machine
- Bioanalyzer®, TapeStation® and associated consumables or other fragment analyzer
Product Notes
- It is possible for FE(II) Solution to have a yellow color. Testing has shown no difference in performance based on color.
Protocols, Manuals & Usage
Protocols
Manuals
Usage & Guidelines
- E7140L_E7120L_EMSeqUDI_IlluminaLRM
- E7140S_E7120S_EMSeqUDI_IlluminaLRM
- Forward Strand Workflow Sample Sheet for E7140L: NovaSeq 6000 with v1.0 reagent kits, MiniSeq with Rapid reagent kits, MiSeq, HiSeq 2000/2500 (paired-end flow cell), HiSeq 3000/4000 (single-read flow cell)
- Forward Strand Workflow Sample Sheet for E7140S: NovaSeq 6000 with v1.0 reagent kits, MiniSeq with Rapid reagent kits, MiSeq, HiSeq 2000/2500 (paired-end flow cell), HiSeq 3000/4000 (single-read flow cell)
- Reverse Complement Workflow Sample Sheet for E7140L: iSeq 100, MiniSeq with Standard reagent kits, NextSeq Systems, NovaSeq 6000 with v1.5 reagent kits, HiSeq 2000/2500 (single-read flow cell), HiSeq 3000/4000 (paired-end flow cell)
- Reverse Complement Workflow Sample Sheet for E7140S: iSeq 100, MiniSeq with Standard reagent kits, NextSeq Systems, NovaSeq 6000 with v1.5 reagent kits, HiSeq 2000/2500 (single-read flow cell), HiSeq 3000/4000 (paired-end flow cell)
FAQs & Troubleshooting
FAQs
- What types of sample can be processed using the NEBNext® Enzymatic Methyl-seq Kit and the Enzymatic Methyl-seq Conversion Module?
- What are the recommended inputs for the NEBNext® Enzymatic Methyl-seq Kit and the Enzymatic Methyl-seq Conversion Module?
- What is the difference between the NEBNext Enzymatic Methyl-seq Kit and the Enzymatic Methyl-seq Conversion Module?
- What buffers are recommended for shearing DNA in NEB’s enzymatic methyl-seq (EM-seq™) workflows?
- What is the concentration of the EM-seq Adaptor and EM-seq Index Primers?
- Can the “active” TET2 Buffer be stored longer than 4 months?
- Can the freshly diluted Fe(II) Solution be stored long term?
- Why, at some stages of the EM-seq protocol, do the NEBNext Sample Purification Beads behave differently when cleaning up the sample?
- What is the expected size of an EM-seq library?
- How should EM-seq libraries be sequenced?
- How should EM-seq sequencing data be analyzed?
- Can the EM-seq Adaptor be substituted with another adaptor?
- Are EM-seq libraries directional or non-directional?
- Can other buffers be used in place of the supplied EM-seq Elution Buffer?
- How are EM-seq libraries prepared from cell-free DNA (cfDNA)?
- Can EM-seq libraries be prepared from FFPE DNA?
- What levels of conversion are typical with the control DNAs supplied in the EM-seq kit?
- Can enzymatically fragmented DNA be used as input material for EM-seq™?
- Are Sample Sheets available for use with the NEBNext® Multiplex Oligos for EM-seq?
- Can I use NaOH (sodium hydroxide) instead of formamide to denature my DNA prior to the deamination reaction?
- Are dual indexed libraries compatible with single end sequencing?
- Is it normal if the FE(II) solution from the EM-seq product is yellow or a color change is observed?
- What percentage of PhiX is needed for EM-seq libraries?
Citations & Technical Literature
Citations
Additional Citations
- Morrison J, et al. (2021) Evaluation of whole-genome DNA methylation sequencing library preparation protocols. Epigenet Chromatin; PREPRINT (version 1) available at Research Square, DOI: 10.21203/rs.3.rs-249202/v1
- Erger F, et al. (2020) cfNOMe - A Single Assay for Comprehensive Epigenetic Analyses of Cell-Free DNA. Genome Med; 12(1):54, PubMedID: 32580754, DOI: 10.1186/s13073-020-00750-5
- Feng S, et al. (2020) Efficient and accurate determination of genome-wide DNA methylation patterns in Arabidopsis with enzymatic methyl sequencing. Epigenet Chromatin; DOI: 10.21203/rs.3.rs-41816/v1
- Sun Z, et al. (2019) Non-destructive enzymatic deamination enables single molecule long read sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single base resolution. bioRxiv; DOI: 10.1101/2019.12.20.885061
- Vaisvila R, et al. (2021) Enzymatic methyl sequencing detects DNA methylation at single-base resolution from picograms of DNA. Genome Res; PubMedID: 34140313, DOI: 10.1101/gr.266551.120
Publications
- Vaisvila R, et al. (2021) Enzymatic methyl sequencing detects DNA methylation at single-base resolution from picograms of DNA. Genome Res; PubMedID: 34140313, DOI: 10.1101/gr.266551.120
Feature Articles
Quality, Safety & Legal
Quality Assurance Statement
Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.Specifications
The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]Certificate Of Analysis
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]- E7120-2_v0_10043692
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Safety DataSheets
The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.Control DNA CpG methylated pUC19
Control DNA CpG unmethylated Lambda
NEBNext® Ultra II End Prep Reaction Buffer
NEBNext® Ultra II End Prep Enzyme Mix
NEBNext® Ultra™ II Ligation Master Mix
NEBNext® Ligation Enhancer
Elution Buffer
TET2 Reaction Buffer
TET2 Reaction Buffer Supplement
Oxidation Supplement
Oxidation Enhancer
TET2
Fe (II) Solution
Stop Reagent
APOBEC
APOBEC Reaction Buffer
BSA
NEBNext® Q5U® Master Mix
DTT
EM-seq™ Index Primer 1 (S)
EM-seq™ Index Primer 2 (S)
EM-seq™ Index Primer 3 (S)
EM-seq™ Index Primer 4 (S)
EM-seq™ Index Primer 5 (S)
EM-seq™ Index Primer 6 (S)
EM-seq™ Index Primer 7 (S)
EM-seq™ Index Primer 8 (S)
EM-seq™ Index Primer 9 (S)
EM-seq™ Index Primer 10 (S)
EM-seq™ Index Primer 11 (S)
EM-seq™ Index Primer 12 (S)
EM-seq™ Index Primer 13 (S)
EM-seq™ Index Primer 14 (S)
EM-seq™ Index Primer 15 (S)
EM-seq™ Index Primer 16 (S)
EM-seq™ Index Primer 17 (S)
EM-seq™ Index Primer 18 (S)
EM-seq™ Index Primer 19 (S)
EM-seq™ Index Primer 20 (S)
EM-seq™ Index Primer 21 (S)
EM-seq™ Index Primer 22 (S)
EM-seq™ Index Primer 23 (S)
EM-seq™ Index Primer 24 (S)
NEBNext® EM-seq™ Adaptor
NEBNext® Sample Purification Beads
NEBNext® 96 Unique Dual Index Primer Pairs Plate (L)
Legal and Disclaimers
Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.
Licenses
In the United States and Europe, this product cannot be used in, or incorporated into, any blood-based, sequencing-based screening diagnostic assay (including product or service-based offerings) for the early detection of colorectal, gastric, lung, liver, ovarian, breast, prostate, esophageal, thyroid, uterine, pancreatic, bladder or kidney cancer indications.
This product is licensed for research and commercial use from Bio-Rad Laboratories, Inc., under U.S. Pat. Nos. 6,627,424, 7,541,170, 7,670,808, 7,666,645, and corresponding patents in other countries. No rights are granted for use of the product for Digital PCR or real-time PCR applications, with the exception of quantification in Next Generation Sequencing workflows.
This product is licensed from University of Newcastle Upon Tyne under EPO Patent No. 1463809B1 and Australian Patent No. AU2003226543.
Nucleic acid-based aptamers for use with thermophilic DNA polymerases are licensed exclusively by New England Biolabs, Inc. from SomaLogic, Inc. New England Biolabs, Inc. gives the Buyer/User a non-exclusive license to use the NEBNext® Enzymatic Methyl-seq Kit for Research Use Only (RUO). Commercial use of this product may require a license from New England Biolabs, Inc. For additional information or to inquire about commercial use, please contact busdev@neb.com.
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The supporting documents available for this product can be downloaded below.