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NEXTFLEX Rapid DNA-Seq Kit 2.0

The NEXTFLEX™ Rapid DNA-Seq Kit 2.0 streamlines whole-genome sequencing (WGS) library preparation from 1 ng to 1 µg of mechanically fragmented DNA, producing Illumina® and Element AVITI™-ready libraries in ~ 3 hours with minimal hands-on time.

An optimized end-repair/A-tail → ligation workflow, followed by a short, high-fidelity PCR step, yields high-complexity libraries with low GC bias and duplication rates, even for GC-rich or degraded samples. The chemistry is automation-ready (validated on Sciclone G3 and Zephyr G3 systems) and supports both single- and paired-end sequencing, making it equally suited for low-input, high-throughput, PCR amplified, or PCR-free WGS projects.

View product information ビュー related reagents

Feature	Specification
Automation Compatible	Yes
Product Group	DNA-seq

View product information ビュー related reagents

Product variants

Unit Size: 8 rxns

Part #:

NOVA-5188-01

Unit Size: 48 rxns

Part #:

NOVA-5188-02

Unit Size: 96 rxns

Part #:

NOVA-5188-03

For research use only. Not for use in diagnostic procedures.

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Product information

Overview

End-to-end WGS library-prep solution including reagents and magnetic beads for optional one- or two-sided size selection
1 ng – 1 µg genomic DNA input; PCR-free libraries possible at ≥ 250 ng
~ 3 h bench-to-sequencer workflow with < 45 min hands-on time
Low GC bias (< 3 %) and low duplication, delivering uniform coverage across GC-rich or moderately degraded genomes
Up to 1,536 unique dual-index (UDI) barcodes available; 96 UDI-UMI sets available for rare-variant detection
Convenient 8, 48, or 96-reaction kit sizes keep cost per sample competitive
Generates single- or paired-end libraries for Illumina® and Element AVITI™ sequencing platforms
Automation-validated on the Sciclone™ G3 NGSx and Zephyr™ G3 NGS systems
Designed for mechanically fragmented DNA (e.g., Covaris®, Bioruptor®); no enzymatic fragmentation module needed
High adapter-ligation efficiency ensures maximum library conversion, even at low DNA inputs.

Additional product information

Higher WGS library yield & conversion across wide input range

The NEXTFLEX Rapid DNA-Seq 2.0 consistently converts a high percentage of mechanically-sheared fragments into adapter-ligated molecules, delivering up to 2 × more final library (nM) than a leading mechanical-ligation kit, even at 1 ng DNA input. The higher conversion translates to a larger pool of unique reads and more uniform coverage in GC-rich regions (≤ 3 % GC-slope).

Library yield and conversion vs competitor: NEXTFLEX Rapid DNA-Seq Kit 2.0 at 250 ng and 1 ng low-input WGS

Figure 1. Superior yield and conversion with NEXTFLEX Rapid DNA-Seq 2.0.
(A) Library yield (nM) from 250 ng and 1 ng inputs of human genomic DNA and E. coli DNA; libraries were amplified with identical PCR-cycle counts. NEXTFLEX Rapid DNA-Seq 2.0 (blue) produces substantially higher yields than Competitor K (purple), especially at 1 ng.
(B) Normalised library-conversion rate (ratio of qPCR-quantified library molecules to input fragment count) for 250 ng human DNA shows.

Reduced GC bias delivers more even genome coverage

GC-coverage analysis shows that NEXTFLEX Rapid DNA-Seq 2.0 preserves uniform read depth from 20% to 80% GC, whereas other mechanical-ligation kits lose reads in GC-rich windows. This uniformity translates to near-complete genome coverage at standard sequencing depths, allowing researchers to call SNPs and assemble contigs with confidence, even in regions that are typically under-represented.

Figure 2: GC-coverage plots show uniform read depth with NEXTFLEX Rapid DNA-Seq 2.0 versus coverage loss in competitor libraries

Greater usable genome coverage at standard sequencing depths

When libraries are sequenced to typical whole-genome depths, NEXTFLEX Rapid DNA-Seq 2.0 delivers a larger proportion of the genome above key coverage thresholds (1X, 5X, 10X) than a leading mechanical-ligation kit. This higher usable coverage, observed in both yeast and E. coli datasets, reduces the need for additional sequencing reads and improves confidence in downstream SNP discovery, structural-variant calling, and de-novo assembly projects.

Bar charts showing genome coverage at 1X, 5X and 10X depth for yeast and E. coli libraries: NEXTFLEX Rapid DNA-Seq 2.0 vs competitor kit

Figure 3: At equivalent read depths, libraries prepared with NEXTFLEX Rapid DNA-Seq 2.0 (blue) cover a larger fraction of the yeast and E. coli genomes at ≥ 1X, ≥ 5X and ≥ 10X thresholds than those prepared with Competitor K (purple). The improved coverage reduces re-sequencing costs and enhances variant-calling accuracy in whole-genome studies.

Flexible Multiplexing Options

The NEXTFLEX Rapid DNA-Seq Kit 2.0 supports high-throughput multiplexing with the full line of NEXTFLEX Adapters offering:

Up to 1,536 unique dual-index pairs (8- or 10-bp) to eliminate index hopping and enable large WGS or capture projects
Single-index and paired-end compatibility for legacy workflows
UDI-UMI plates (96 pairs) that add unique molecular identifiers for accurate low-frequency variant detection and duplex sequencing
Color-balanced index sets optimized for Illumina® patterned flow-cells and Element AVITI™ chemistry

Automation Compatibility

The NEXTFLEX Rapid DNA-seq Kit 2.0 is compatible with multiple liquid handlers, including the Revvity Sciclone G3 NGSx and Zephyr G3 NGS workstations.

Specifications

Other specifications

Automation Compatible	Yes
Product Group	DNA-seq
Shipping Conditions	Dual Temperature
Unit Size	8 rxns

Citations

Ferrari, G., Esselens, L., Hart, M. L., Janssens, S., Kidner, C., … & von Rintelen, T. (2023). Developing the protocol infrastructure for DNA sequencing natural history collections. Biodiversity Data Journal, 11, e102317. https://doi.org/10.3897/BDJ.11.e102317
Gaulke, C. A., Schmeltzer, E. R., Dasenko, M., Tyler, B. M., Vega Thurber, R., & Sharpton, T. J. (2021). Evaluation of the effects of library preparation procedure and sample characteristics on the accuracy of metagenomic profiles. mSystems, 6(5), e00440-21. https://doi.org/10.1128/mSystems.00440-21
Lando, D., Ma, X., Cao, Y., … & Laue, E. D. (2024). Enhancer-promoter interactions are reconfigured through the formation of long-range multiway hubs as mouse ES cells exit pluripotency. Molecular Cell, 84(8), 1406–1421.e8. https://doi.org/10.1016/j.molcel.2024.02.015
Ma, Y., Schwager, A., Dib, C., … & Ferrer, M. (2024). Exchange of subtelomeric regions between chromosomes 4q and 10q reverts the FSHD genotype and phenotype. Science Advances, 10(18), eadl1922. https://doi.org/10.1126/sciadv.adl1922

FAQs

What makes this kit better for low-input whole-genome sequencing than other library-prep kits?

NEXTFLEX Rapid DNA-Seq 2.0 uses a combined end-repair/A-tail chemistry and high-efficiency ligase that converts a high percentage of fragments into adapter-ligated molecules. Because so little DNA is lost in the process, you can start with as little as 1 ng of fragmented DNA and still obtain complex libraries with low duplication rates and GC bias - performance most kits only achieve at ≥ 10 – 50 ng.
Do I need a Covaris® instrument, or can I use the kit with enzymatic fragmentation?

The kit is designed for mechanical fragmentation (e.g., Covaris, Bioruptor, Megaruptor). You can use DNA that was previously sheared enzymatically, but the workflow itself does not include a fragmentation step. If you need an all-in-one WGS library prep plus enzymatic fragmentation solution, consider the NEXTFLEX Rapid XP V2 kit.
Can I run PCR-free libraries with this kit?

Yes, if you have ≥ 250 ng of high-quality genomic DNA. Skip the PCR step after ligation, perform a standard bead cleanup, and proceed directly to QC and pooling. PCR-free libraries benefit variant-calling applications by reducing amplification artefacts and GC bias even further.
How does the kit handle GC-rich genomes or degraded samples?

Mechanical shearing plus bias-mitigated ligation chemistry yields a flat GC-coverage profile (≤ 3% slope). Internal data show near-complete coverage on a 68%-GC Bordetella genome and successful library prep from moderately degraded FFPE DNA without protocol changes - so GC-rich or damaged templates sequence reliably.
Can I combine these libraries with target-capture panels?

Yes. Libraries generated with NEXTFLEX Rapid DNA-Seq 2.0 have standard Illumina® P5/P7 ends and are fully compatible with Twist®, Agilent® SureSelect, IDT® xGen, and other hybrid-capture or exome-capture workflows. Shear to ~200–250 bp if your capture panel specifies shorter inserts, then proceed with the vendor’s normal hybridization and post-capture PCR protocol.
How should I store the kit and what is the shelf life?
- Enzyme mixes & buffers: store at –20°C; stable for at least 6 months from date of receipt.
- NEXTFLEX Cleanup Beads 2.0: store at 4°C; warm to room temperature and vortex before use.
- Resuspension Buffer & nuclease-free water: 4°C or room temperature.
- The kit ships on dry ice; avoid > 3 freeze-thaw cycles to maintain ligase activity.

Which sample types has the kit been proven to work with?

Sample type	Typical input	Notes
High-quality gDNA (human, plant)	5 – 250 ng	Standard WGS; PCR-free at ≥ 250 ng
FFPE or mildly degraded DNA	10 – 100 ng	Skip size-selection to maximize yield
Microbial & metagenomic DNA	1 – 50 ng	Handles broad GC range (20 – 80% GC)
Low-mass environmental samples	1 – 10 ng	Works with Covaris-sheared eDNA or amplicons