ONT Native Barcoding Kit 96 V14

The Native Barcoding Kit 96 V14 is a comprehensive solution for PCR-free multiplexing of up to 96 DNA samples, designed to achieve high accuracy and maintain the integrity of base modifications, critical for epigenetic studies and long-read sequencing.

Key Features and Benefits

1. Multiplexing Capability:
   • Supports up to 96 samples per flow cell, significantly reducing the cost per sample.
   • Optimized for experiments requiring high data efficiency when sequencing smaller data per sample.
2. High Accuracy:
   • Modal raw read accuracy of Q20+ (≥99%) when used with the R10.4.1 nanopore.
   • Compatibility with Kit 14 sequencing chemistry ensures precise results.
3. PCR-Free Workflow:
   • Preserves base modifications and avoids PCR-related bias.
   • Allows retention of native DNA properties.
4. Flexible Read Length:
   • Enables control over read length, suitable for studies requiring long reads (e.g., structural variant analysis).
5. Upstream Compatibility:
   • Supports whole-genome amplification and size selection for tailored workflows.
   • Compatible with ONT’s Short Fragment Eliminator Kit (EXP-SFE001) for size selection.
6. Ease of Use:
   • Integrated workflow similar to the Ligation Sequencing Kit protocol.
   • Deconvolution supported by MinKNOW, Guppy, and EPI2ME software.
7. Optimized Reagents:
   • Includes updated adapters and buffers for improved sequencing performance and reduced EDTA concentrations for enhanced stability.

Specifications

Workflow

The preparation involves the following steps:

1. DNA Repair and dA-Tailing:
   • Genomic DNA undergoes repair and dA-tailing using the NEBNext End Repair/dA-Tailing module.
   • DNA repair skipped if amplicons are used.
2. Barcode Adapter Ligation:
   • Unique dT-tailed barcode adapters are ligated to the dA-tailed DNA.
   • Barcode adapters also include a cohesive end that binds the sequencing adapter.
3. Pooling:
   • Barcoded samples are pooled, reducing handling time and costs.
4. Sequencing Preparation:
   • Samples are prepared using ONT’s proprietary reagents and loaded onto the flow cell.
5. Data Analysis:
   • MinKNOW, Guppy, and EPI2ME tools classify and sort reads by barcodes.

What’s Included in the Kit

Third-Party Materials and Equipment Required

Reagents:

• NEB Blunt/TA Ligase Master Mix (M0367).
• NEBNext FFPE Repair Mix (M6630).
• NEBNext Ultra II End Repair/dA-Tailing Module (E7546).
• AMPure XP Beads.
• Qubit dsDNA HS Assay Kit and Nuclease-Free Water.

Equipment:

• Thermal cycler, magnetic rack, microfuge, Hula mixer, vortex mixer.
• Multichannel pipettes and standard laboratory consumables.

Shipping and Logistics

• Environmentally friendly, temperature-controlled shipping.
• Products shipped Monday–Thursday (varies by region).

Advantages

1. Cost Efficiency:
   • Significantly reduces price per sample when pooling 96 samples.
2. Adaptability:
   • Compatible with various input types, including low-concentration DNA.
3. Accuracy and Data Quality:
   • Achieves high-accuracy sequencing with R10.4.1 flow cells.
   • Maintains native DNA properties for epigenetic and long-read studies.

The Native Barcoding Kit 96 V14 is a versatile tool designed for researchers seeking cost-effective, high-accuracy sequencing with full control over sample properties and read length, making it suitable for a variety of applications, from genomic studies to pathogen surveillance.