NEXTFLEX® Rapid DNA-Seq Library Prep Kit for Illumina® Platforms

  • Flexible amounts of input DNA from 1 ng to 1 µg
  • Fast workflow
  • Flexible barcode options – up to 384 single-index barcodes available
  • Compatible with Illumina® sequencing platforms including the MiSeq®, HiSeq®, NextSeq® and HiSeq® X Ten

We recommend the new NEXTFLEX® Rapid DNA-Seq Kit 2.0 for your DNA-seq library prep needs.

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We have a new improved DNA-seq library prep kit,
NEXTFLEX® Rapid DNA-Seq Kit 2.0.

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

  • NOVA-5144-01


    8 RXNS

  • NOVA-5144-02


    48 RXNS

  • NOVA-5144-03


    BARCODES 1-24
    48 RXNS

  • NOVA-5144-04


    BARCODES 25-48
    48 RXNS

Rapid DNA Library Prep for Sequencing on Illumina® Platforms

The NEXTFLEX® Rapid DNA-Seq Kit is a rapid DNA library prep kit producing libraries for Illumina® sequencing from as little as 1 ng of DNA in two hours. This kit is ideal for use with genomic DNA, FFPE samples, ChIP DNA, upstream target capture samples, pooled amplicons and low input clinical samples. This DNA-Seq library prep kit is highly flexible in terms of sample requirements, accommodating a range of input DNA amounts from 1 ng to 1 µg. The optimized protocol allows for fast generation of non-size-selected libraries and size-selected libraries using reproducible gel-free double SPRI selection.

Rapid DNA-Seq Protocol

Flexible Multiplexing Options

The NEXTFLEX® Adapters are long, annealed adapters containing indexed sequences that offer an improved multiplexing workflow and flexible setup. These barcodes can be used with single, paired-end and multiplex reads. We strives to make available flexible barcoding options. The NEXTFLEX® Rapid DNA-Seq Kit is designed to be used with the NEXTFLEX® Unique Dual Index barcodes, NEXTFLEX® DNA barcodesNEXTFLEX-96 DNA barcodes and NEXTFLEX-HT barcodes when 10 ng or more of starting material is used. These barcodes are available in sets of 6, 12, 24, 48, 96 and 384 unique indexed adapters.

When less than 10 ng of starting material is available, the NEXTFLEX® ChIP-Seq barcodes and NEXTFLEX-96 ChIP-Seq barcodes should be used. These barcodes are available in sets of 6, 12, 24, 48, and 96 unique indexed adapters.

Cost Effective NEXTFLEX® Rapid DNA-Seq Kit Bundles

To simplify your DNA sequencing PerkinElmer offers affordable bundles containing NEXTFLEX® Rapid DNA-Seq library prep reagents and NEXTFLEX® DNA barcodes. There are two different versions of this kit. They each contain the 48 rxn NEXTFLEX® Rapid DNA-Seq Kit and 24 unique NEXTFLEX® DNA Barcodes (in aliquots of 2 reactions each).

Automation Compatibility

The NEXTFLEX® Rapid DNA-Seq Library Prep Kit was designed for easy migration onto automated liquid handling platforms. A validated automation protocol for use with the PerkinElmer Sciclone® G3 NGS and NGSx workstations is available.


  • NEXTFLEX® End-Repair & Adenylation Buffer Mix
  • NEXTFLEX® End-Repair & Adenylation Enzyme Mix
  • NEXTFLEX® Ligase Enzyme Mix
  • NEXTFLEX® PCR Master Mix
  • Nuclease-free Water
  • NEXTFLEX® Sizing Solution
  • NEXTFLEX® Resuspension Buffer


  • 1 ng – 1 µg of fragmented DNA in up to 32 µL nuclease-free water.
  • NEXTFLEX® DNA Barcodes – 6 / 12 / 24 / 48 (Cat # 514101, 514102, 514103, 514104) or NEXTFLEX-96 DNA Barcodes (Cat # 514106)
  • Ethanol 100% (room temperature)
  • Ethanol 80% (room temperature)
  • 96 well PCR Plate Non-skirted (Phenix Research®, Cat # MPS-499) or similar
  • 96 well Library Storage and Pooling Plate (Fisher Scientific®, Cat # AB-0765) or similar
  • Adhesive PCR Plate Seal (BioRad, Cat # MSB1001)
  • Agencourt® AMPure® XP 60 mL (Beckman Coulter Genomics®, Cat # A63880)
  • Magnetic Stand -96 (Thermo Fisher® Scientific, Cat # AM10027) / or / similar
  • Thermocycler
  • 2, 10, 20, 200 and 1000 µL pipettes / multichannel pipettes
  • Nuclease-free barrier pipette tips
  • Vortex



  • Rapid DNA-Seq Kit 2.0 (2018)


  • Rapid DNA-Seq Kit 2.0 (2018)

Publications that Cite Using the NEXTFLEX® Rapid DNA-Seq Kit:

  • Brodie, J., et al. (2016) Characterising the microbiome of Corallina officinalis, a dominant calcified intertidal red alga. FEMS Microbiology Ecology. doi: 10.1093/femsec/fiw110.
  • Campana, M. G., et al. (2014) False positives complicate ancient pathogen identifications using high-throughput shotgun sequencing. BMC Research Notes. 7:111 http://www.biomedcentral.com/1756-0500/7/111.
  • Campbella, M. A., et al. (2015) Genome expansion via lineage splitting and genome reduction in the cicada endosymbiont Hodgkinia. PNAS. doi: 10.1073/pnas.1421386112.
  • Cervera-Juanes, R., et al. (2015) MAOA expression predicts vulnerability for alcohol use. Molecular Psychiatry. doi:10.1038/mp.2015.93.
  • Chwialkowska, K., Nowakowska, U., Mroziewicz, A., Szarejko, I., and Kwasniewski, M. (2016) Water-deficiency conditions differently modulate the methylome of roots and leaves in barley (Hordeum vulgare L.) J. Exp. Bio. doi: 10.1093/jxb/erv552.
  • Devall, M., et al. (2015) A comparison of mitochondrial DNA isolation methods in frozen post-mortem human brain tissue—applications for studies of mitochondrial genetics in brain disorders. Biotechniques 59(4): 241-246.
  • DeWitt, M., Magis, W., Bray, N. et al. (2016, Jan 15). Efficient Correction of the Sickle Mutation in Human Hematopoietic Stem Cells Using a Cas9 Ribonucleoprotein Complex Retrieved from http://biorxiv.org/content/early/2016/01/15/036236
    doi: 10.1101/036236
  • Gal, C., et al. (2015) Abo1, a conserved bromodomain AAA‐ATPase, maintains global nucleosome occupancy and organization. EMBO Reports. 16:11. pp 1409 – 1580. doi: 10.15252/embr.201540476.
  • Gal, C. et. al. (2015) The impact of the HIRA histone chaperone upon global nucleosome architecture. Cell Cycle. 14:1, 123-134. doi:10.4161/15384101.2014.967123.
  • Gan, H. M., Tan, M. H., Lee, Y. P. and Austin, C. M. (2014) The complete mitogenome of the Australian tadpole shrimp Triops australiensis (Spencer & Hall, 1895) (Crustacea: Branchiopoda: Notostraca).
  • Gan, H. M., Tan, M. H., Lee, Y. P. and Austin, C. M. (2014) The complete mitogenome of the river blackfish, Gadopsis marmoratus (Richardson, 1848) (Teleostei: Percichthyidae). Mitochondrial DNA. doi:10.3109/19401736.2014.974174.
  • Givnish, T. J., et al. (2016) Phylogenomics and historical biogeography of the monocot order Liliales: out of Australia and through Antarctica. Cladistics. doi: 10.1111/cla.12153.
  • Goh, H. F. and Philip, K. (2015) Purification and Characterization of Bacteriocin Produced by Weissella confusa A3 of Dairy Origin. PLoS ONE. doi: 10.1371/journal.pone.0140434.
  • Gultekin, S. E., et al. (2016) Unusual Presentation of an Adenocarcinoma of the Lung Metastasizing to the Mandible, Including Molecular Analysis and a Review of the Literature. Journal of Oral and Maxillofacial Surgery, doi:10.1016/j.joms.2016.06.004.
  • Harrisson, K., et al. (2016) Pleistocene divergence across a mountain range and the influence of selection on mitogenome evolution in threatened Australian freshwater cod species. Heredity. 1–10. doi:10.1038/hdy.2016.8.
  • Hill, C. J. et al. (2016) Effect of room temperature transport vials on DNA quality and phylogenetic composition of faecal microbiota of elderly adults and infants. Microbiome. 4:19. doi: 10.1186/s40168-016-0164-3.
  • Laver, T., Harrison, J., O’Neill, P. A., Moore, K., Farbos, A., Paszkiewicz, K. and Studholme, D. J. (2015) Assessing the performance of the Oxford Nanopore Technologies MinION. Biomolecular Detection and Quantification 3 (2015) 1–8. doi :10.1016/j.bdq.2015.02.001.
  • Palomo, A., et al. (2016) Metagenomic analysis of rapid gravity sand filter microbial communities suggests novel physiology of Nitrospira spp. The ISME Journal. doi:10.1038/ismej.2016.63.
  • Shain, A. H., et al. (2015) The Genetic Evolution of Melanoma from Precursor Lesions. The New England Journal of Medicine. 373:1926-1936. doi: 10.1056/NEJMoa1502583.
  • Tatarenkova, A., Mesaka F. and Avisea, J. C. (2015) Complete mitochondrial genome of a self-fertilizing fish Kryptolebias marmoratus (Cyprinodontiformes, Rivulidae) from Florida. Mitochondrial DNA. doi: 10.3109/19401736.2015.1115861.
  • Yang, Y. A., et al. (2016) FOXA1 potentiates lineage-specific enhancer activation through modulating TET1 expression and function. Nucleic Acids Research. doi: 10.1093/nar/gkw498.

When to Size Select
Size selection is a critical issue in NGS library preparation. Our blog post, Tech Tips – When to Size Select, addresses a number of factors to consider before determining a size selection strategy.

The NEXTFLEX®  Rapid DNA-Seq Kit contains enough material to prepare 8 or 48 DNA samples for Illumina® sequencing. The shelf life of all reagents is at least 12 months when stored properly. All components can be safely stored at -20°C. These kits ship on dry ice.