Product Class: Other

NEBNext® FFPE DNA Repair Mix

For superior performance, the NEBNext FFPE DNA Repair v2 Module (NEB #E7360) is now available.

Kits for FFPE DNA library prep are now available, with or without enzymatic DNA fragmentation (NEBNext UltraShear™ FFPE DNA Library Prep Kit NEB #E6655 and NEBNext FFPE DNA Library Prep Kit NEB #E6650).

Product Introduction

Archiving of clinical materials as Formalin-Fixed, Paraffin-Embedded (FFPE) samples is a common practice. However, the methods used for fixation and storage significantly damage and compromise the quality of DNA from these samples. As a result, it can be challenging to obtain useful information, including high-quality sequence data, especially when sample amounts are limited.
 
The NEBNext FFPE DNA Repair Mix is a cocktail of enzymes formulated to repair DNA, and specifically optimized and validated for repair of FFPE DNA samples.
 
Catalog # Size Concentration
M6630S 24.0 reactions
M6630L 96.0 reactions

Product Information

Description



The NEBNext FFPE DNA Repair Mix contains enzymes and buffers that are optimized to repair FFPE DNA in next generation sequencing workflow. The mix can be used upstream of any next generation sequencing library construction method, for any sequencing platform.
 

Table 1: Types of FFPE DNA damage and their ability to be repaired by the NEBNext FFPE DNA Repair Mix
FFPE DAMAGE TYPE REPAIRED BY THE FFPE DNA ENZYME REPAIR MIX?
Deamination of cytosine to uracil Yes
Nicks and gaps Yes
Oxidized bases Yes
Blocked 3´ ends Yes
DNA fragmentation No
DNA-protein crosslinks No

Figure 1: FFPE DNA repair increases NGS library yields
An example of Agilent Bioanalyzer® traces of libraries prepared from stomach tumor FFPE DNA that was treated with the FFPE DNA Repair Mix, or was untreated, before library construction. Yield improvements of 10% to 458% have been observed. 

Figure 2. FFPE DNA repair increases NGS library yields 
FFPE DNA samples (25 ng or 50 ng) from a variety of tissue types were treated with the NEBNext FFPE DNA Repair Mix, or were untreated, before library construction. Library yields were measured using the Agilent Bioanalyzer, and ratios of library yields with and without treatment with the NEBNext FFPE DNA Repair Mix were calculated. Yield improvements with these samples ranged from 101% to 458%. 

Figure 3. FFPE DNA Repair improves sequence quality from FFPE samples and does not introduce new mutations 
Genomic DNA was extracted using standard DNA purification. Each DNA sample was treated with the NEBNext FFPE DNA Repair Mix, or was untreated, before library construction and Illumina® sequencing.
Panel A: The effect of DNA repair on the percentage of mappable reads. Reads were mapped to the human genome (hg19) using BWA (1).
Panel B: The effect of DNA repair on the percentage of reads that were properly paired from a paired-end sequencing protocol.

Lot Control

The lots provided are managed separately and qualified by additional functional validation. Individual reagents undergo standard enzyme activity and quality control assays, and also meet stringent criteria in the additional quality controls listed on each individual component page
This product is related to the following categories:
FFPE DNA Products,
Next Generation Sequencing Library Preparation Products,

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  • M6630S     -20    
      NEBNext® FFPE DNA Repair Mix M6630SVIAL -20 1 x 0.048 ml Not Applicable
      NEBNext FFPE DNA Repair Buffer E6622AVIAL -20 1 x 0.156 ml Not Applicable
  • M6630L     -20    
      NEBNext® FFPE DNA Repair Mix M6630LVIAL -20 1 x 0.192 ml Not Applicable
      NEBNext FFPE DNA Repair Buffer E6622AAVIAL -20 1 x 0.624 ml Not Applicable

Properties & Usage

Materials Required but not Supplied

Library Construction Reagents (e.g., NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB #E7645))

  • 100% Ethanol
  • Nuclease-free Water
  • 10 mM Tris-HCl, pH 8.0 or 0.1X TE
  • DNA LoBind Tubes (Eppendorf #022431021)
  • AMPure® XP Beads (Beckman Coulter, Inc. #A63881)
  • NEBNext Multiplex Oligos for Illumina
  • Magnetic rack or plate (e.g., NEBNext® Magnetic Separation Rack (NEB #S1515S), Alpaqua ® 96S Super Magnet Plate (#A001322), or equivalent)
  • PCR Machine

Protocols, Manuals & Usage

Protocols

  1. Where can I find guidelines and protocols for using FFPE DNA with NEBNext Products?

Manuals

The Product Manual includes details for how to use the product, as well as details of its formulation and quality controls.

FAQs & Troubleshooting

FAQs

  1. Will the FFPE DNA Repair Mix ligate my DNA fragments?
  2. Will treating my DNA with the FFPE DNA Repair Mix hurt my downsteam reaction?
  3. Will the FFPE DNA Repair Mix blunt the ends of the DNA?
  4. Does the FFPE DNA Repair Mix insert random nucleotides into the sequence that it repairs?
  5. Does the FFPE DNA Repair Mix remove covalent modifications from DNA bases, such as methylation, biotin or digoxigenin?
  6. Does the FFPE DNA Repair Mix repair DNA-protein crosslinks?
  7. Does the FFPE DNA Repair Mix fix blocked 3’ ends?
  8. Can the FFPE DNA Repair Mix repair damage in both single and double stranded DNA? Or, does it require double stranded DNA as a template?
  9. If I had a DNA template with mutation sites (ie. 8-oxoguanine or deaminated cytosines) that are directly adjacent to each other on opposite strands would treatment with FFPE DNA Repair Mix cause a double strand nick/break?
  10. What gap lengths can be repaired with the FFPE DNA Repair Mix?
  11. When working with fragmented DNA, will the ends be ligated together by the FFPE DNA Repair Mix Reaction Mix?
  12. How are abasic sites repaired by the FFPE DNA Repair Mix Reaction Mix?
  13. Can I use the FFPE DNA Repair Mix to repair other types of low quality DNA other than FFPE DNA?
  14. What is the difference between the NEBNext FFPE DNA Repair v2 Module (NEB #E7360) and the NEBNext FFPE DNA Repair Mix (NEB #M6630)?

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]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

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.

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