NEBNext® Ultra™ II DNA Library Prep

Are you challenged with trying to obtain higher library yields using ever-decreasing input amounts? Each component in the NEBNext Ultra II DNA Library Prep Kit from NEB® has been reformulated, resulting in a several-fold increase in library yield with as little as 500 picograms of input DNA. These advances deliver unprecedented performance, while enabling lower inputs and fewer PCR cycles.

Now available: NEBNext Ultra II DNA Library Prep with Sample Purification Beads, supplied with SPRIselect® beads for size selection and clean-up steps. 

Get even more from less with NEBNext Ultra II – Request your sample today!

Advantages of the NEBNext Ultra II DNA Library Prep Kit for Illumina®

  • Get more of what you need, with the highest library yields
  • Use to generate high quality libraries even when you have only limited amounts of DNA, with inputs as low as 500 pg
  • Prepare libraries from ALL of your samples, including GC-rich targets and FFPE DNA samples
  • Improve yields for target enrichment applications
  • Save time with streamlined workflows, reduced hands-on time, and automation compatibility, and enjoy the flexibility of kit or module format products
  • Enjoy the flexibility and reliability of the gold standard SPRIselect size selection and clean-up beads, supplied in just the amounts you need with the latest version of the kit
  1. NEBNext Ultra II DNA Library Prep Protocol

    This video walks you through DNA Library Preparation using the NEBNext Ultra II DNA Library Prep Kit. The video also includes tips for optimization as well as safe stopping points.

  2. Size Selection and Cleanup with NEBNext Ultra II and SPRI beads

    This video walks you through the size selection and cleanup steps using the NEBNext Ultra II DNA

  3. 12 Quick Tips for NGS Library Preparation

    Using beads to clean up your DNA prior to NGS library prep can be quick and easy, if you follow these few, simple tips.

See what others are saying about Ultra II

“Approximately 88% of the fragments had sequencing adaptors after ligation, far higher than most other library prep kits.“

- Major UK sequencing center

"The new NEBNext DNA Ultra II kit has provided us with a critical opportunity to process challenging low-input genomic samples. These samples otherwise didn’t yield libraries of adequate complexity required for exploring their genomes comprehensively. In libraries prepared with a different kit, the proportion of duplicated reads, which don’t provide new information, is too high. NEBNext DNA Ultra II has alleviated this problem and enabled us to achieve high quality, high content sequencing data that are relevant for our users.“

– Vladimir Benes, Ph.D., Head, GeneCore Facility, EMBL Heidelberg, Germany

"The protocol is very easy to follow and is a step up from the standard NEB Ultra Kit. Using as little as 500 pg of cell-free DNA input, we were able to generate libraries that were successfully used for subsequent exome capture.”

- Christopher Smith, Ph.D., Post-Doctoral Research Associate at the Cancer Research UK - Cambridge Institute (CRUK-CI)

NEBNext Ultra II produces the highest yield libraries from a broad range of input amounts.

Libraries were prepared from Human NA19240 genomic DNA using the input amounts and numbers of PCR cycles shown. Manufacturers’ recommendations were followed, with the exception that size selection was omitted.

NEBNext Ultra II produces the highest rates of conversion to adaptor-ligated molecules from a broad range of input amounts.

Libraries were prepared from Human NA19240 genomic DNA using the input amounts and library prep kits shown without an amplification step, and following manufacturers’ recommendations. qPCR was used to quantitate adaptor-ligated molecules, and quantitation values were then normalized to the conversion rate for Ultra II. The Ultra II kit produces the highest rate of conversion to adaptor-ligated molecules, for a broad range of input amounts.

Read depth correlation shows consistently high coverage for 500 pg–100 ng input amounts.

Libraries were prepared with 100 ng, 1 ng and 500 pg of human NA19240 genomic DNA. Each library was downsampled (sambamba view -s) to include 423 M reads and mapped to GRCh37 using Bowtie 2.2.4. Coverage of each 10 kb region of GRCh37 (as determined by bedtools coverage) was compared between low (500 pg and 1 μg) and 100 ng input. Most regions are covered by ~1,000 reads, as expected. Low and high coverage regions are well correlated.