NEBNext ULTRAEXPRESS® DNA Library Prep Kit

Catalog #	Concentration	Size	List Price	Quantity	Your Price
E3325L		96 reactions	$3,078.00		$2,770.20
E3325S		24 reactions	$809.00		$728.10

Catalog #	Size	List Price	Your Price
E3325L	96 reactions	$3,078.00	$2,770.20
E3325S	24 reactions	$809.00	$728.10

View or download extensive performance data below and in our Data Supplement.

Responding to user need for a faster, streamlined DNA prep workflow that delivers high quality libraries from a variety of sample types, the NEBNext UltraExpress DNA Library Prep Kit has a single protocol for all DNA inputs (10 – 200 ng of pre-sheared DNA). The workflow incorporates master mixed reagents, reduced incubation times, and fewer cleanup steps. Use of this kit also generates less plastic consumable waste because the entire library prep is conducted in a single tube.

Go from pre-sheared sample to library in under 2 hours
Single-protocol simplicity cuts down on reaction setup time, while streamlined workflows speed up library prep
A single-tube workflow means less plastic/consumable waste
Flexibility is enabled with simple guidelines for customized protocols, if desired
Automation-friendly protocols for enhanced scalability

Graph of Library Yield — Libraries were prepared from 10, 50, 100 or 200 ng of Human NA19240 genomic DNA (Coriell Institute for Medical Research) using the same adaptor amount and 8 PCR cycles. Yields exceeded the minimum requirement (40 ng) for a single Ilumina^® NovaSeq^® 6000 run to achieve whole genome sequencing with at least 30X coverage.

Graphed Input Amounts — Libraries were prepared from 10, 50, 100 or 200 ng of Human NA19240 genomic DNA (Coriell Institute for Medical Research) using the same adaptor amount and 8 PCR cycles. Libraries were pooled and sequenced on an Illumina^® MiSeq^® (2 x 75 bases). Data showed consistent GC coverage (A) and insert size (B). 2 million paired-end reads from each library were sampled (seqtk v1.3), adapter-trimmed (seqprep v0.1) and mapped to the GRCh38 reference (bowtie2 v2.4.5), and GC coverage information was calculated using Picard's CollectGCBiasMetrics (v 1.56.0). In (A), the horizontal grey line indicates the expected normalized coverage of 1.0, and the dots in shades of green represent read numbers at each GC%. The grey area plot is a histogram representing the distribution of GC content in 100 bp windows of the reference genome.

Graph of Sample Type — Libraries were prepared using a single protocol from cell-free DNA (cfDNA, 12 ng) without shearing and Maize DNA (10 ng and 100 ng) sheared to 200bp (Covaris^® ME220). Libraries were pooled and sequenced on an Illumina^® MiSeq^® (2 x 75 bases). 1.4 million paired-end reads from each library were sampled (seqtk v1.3), adapter-trimmed (seqprep v0.1), and aligned to either the GRCh38 reference (human) or Zea mays reference genome (maize) (bowtie2 v2.4.5).

A. All sample types showed representative GC coverage. High- and low-input Maize DNA generated consistent GC coverage. The horizontal grey line indicates the expected normalized coverage of 1.0, and the colored dots represent read numbers at each GC%. The grey area plot is a histogram representing the distribution of GC content in 100 bp windows of the reference genome.

B. cfDNA insert size had the characteristic fragmentation pattern with 167 bp peak size and periodicity feature of nucleosome-bound DNA in shorter fragments. 10 ng and 100 ng Maize DNA showed consistent insert size, as expected with the shearing protocol used.

Graphs of library complexity — Libraries were prepared using a single protocol from (A) 10 ng and 100 ng of the ZymoBIOMICS^® Microbial Community DNA Standard (Zymo Research^®, Catalog #D6306), and (B) 10 ng and 100 ng of the ZymoBIOMICS Microbial Community DNA Standard II (Log Distribution) (Zymo Research, Catalog #D6311). Libraries were pooled and sequenced on an Illumina^® MiSeq^® (2 x 75 bases). 1.4 million paired-end reads from each library were sampled (seqtk v1.3), adapter-trimmed (seqprep v0.1) and aligned to a composite reference genome (bowtie2 v2.4.5). 1,000 bp windows of constituent genomes were counted (bedtools 2.30.0) and compared across replicates and input levels. High correlation was observed between replicates and between inputs.

Graph of Complex Mixtures — Libraries were prepared using a single protocol from 10 ng and 100 ng of the ZymoBIOMICS Microbial Community DNA Standard II (Log Distribution) (Zymo Research^®, Catalog # D6311). Libraries were pooled and sequenced on an Illumina^® MiSeq^® (2 x 75 bases). 1.4 million reads from each library were sampled (seqtk v1.3), adapter-trimmed (seqprep v0.1), and aligned to a composite reference genome (bowtie2 v2.4.5). 1,000 bp windows of constituent genomes were counted (bedtools 2.30.0) and compared across expected and detected composition for both input levels. Consistent correlation between expected and detected composition was noted (R2 = 0.96 for 10 ng and R2 = 0.97 for 100 ng). Expected composition: Listeria monocytogenes 89.1%, Pseudomonas aeruginosa 8.9%, Bacillus subtilis 0.89%, Saccharomyces cerevisiae 0.89%, Escherichia coli 0.089%, Salmonella enterica 0.089%, Lactobacillus fermentum 0.0089%, Enterococcus faecalis 0.00089%, Cryptococcus neoformans 0.00089% and Staphylococcus aureus 0.000089%.

Reagents Supplied

The following reagents are supplied with this product:

NEB #

Component Name

Component #

Stored at (°C)

Amount

Concentration

E3325S -20

NEBNext UltraExpress^® End Prep Reaction Buffer	E3324AVIAL	-20	1 x 0.056 ml
NEBNext UltraExpress^® End Prep Enzyme Mix	E3326AVIAL	-20	1 x 0.024 ml
NEBNext UltraExpress^® Ligation Master Mix	E3327AVIAL	-20	1 x 0.24 ml
NEBNext^® MSTC™ High Yield Master Mix	E3328AVIAL	-20	1 x 0.96 ml
NEBNext^® Bead Reconstitution Buffer	E3339AVIAL	-20	1 x 0.096 ml
0.1X TE	E3341AVIAL	-20	1 x 2.4 ml

E3325L -20

NEBNext UltraExpress^® End Prep Reaction Buffer	E3324AAVIAL	-20	1 x 0.221 ml
NEBNext UltraExpress^® End Prep Enzyme Mix	E3326AAVIAL	-20	1 x 0.096 ml
NEBNext UltraExpress^® Ligation Master Mix	E3327AAVIAL	-20	1 x 0.96 ml
NEBNext^® MSTC™ High Yield Master Mix	E3328AAVIAL	-20	1 x 3.84 ml
NEBNext^® Bead Reconstitution Buffer	E3339AAVIAL	-20	1 x 3.84 ml
0.1X TE	E3341AAVIAL	-20	1 x 9.6 ml

Properties & Usage

Companion Products

FAQs

Quality Control Assay

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]

E3325S_L_G_v1

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]

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