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).
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.
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
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