Twist FlexPrep UHT Library Preparation Kit

Simplified Workflow for Ultra-High Throughput Sequencing

Twist’s FlexPrep^TM UHT Library Preparation Kit offers an NGS alternative to traditional microarray-based technologies for population studies, agrigenomics and other ultra-high throughput applications. Combining Twist’s Normalization by Ligation^TM (NBL) technology with enzymatic fragmentation methods, the kit offers built-in sample normalization across a wide range of DNA inputs, reducing both cost and complexity of typical sample processing steps. Early sample barcoding enables downstream pooling of twelve separate samples into one reaction for a more streamlined and efficient workflow.

PRODUCT HIGHLIGHT

Normalization by Ligation (NBL)

Eliminate upfront normalization

No need for intermediate quantitation

Work with sample batches using a wide range of inputs (30 ng to 300 ng)

Early Sample Pooling Saves Cost

Pool 96 samples into 8 tubes after the ligation reaction

Miniaturized reaction volumes upstream

Reduction in consumables downstream

Designed for Ultra-High Capacity Multiplexing

Remove multiplexing limitation per flow cell lane with ultra-high throughput indexing

Increase capacity 12x with existing lab setup*

Supports 96 samples per target enrichment reaction

For research use only. Not for use in any diagnostic or clinical procedures.
*Based on a 8 well vs 96 well workflow

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FAQ

What is Normalization by Ligation?

What sample types and species demo data are available for download?

What are the recommendations for DNA input and quantity?

What are the recommendations for DNA quality?

Can I prepare less than 192 samples at one time?

Integrated Normalization Eliminates Steps

The Twist FlexPrep^TM UHT Library Preparation Kit takes a novel Normalization by Ligation^TM approach which eliminates the need for upfront and intermediate sample quantitation, streamlining your sequencing workflow.

Simplicity of Workflow

Fragmentation and ligation reactions are prepared in one well per sample. The adapters used during ligation contain inline barcodes that allow for the pooling of all 12 wells in a row of a 96-well plate. Individual pools are prepared with indices (UDIs) added by PCR for pool-level demultiplexing. Sequencing throughput can be maximized by running up to 1,152 samples in a single sequencing run from one FlexPrep kit. This increased efficiency can translate to cost and consumables savings.

Normalized Yield with FlexPrep UHT*

NGS read depth normalization with variable DNA mass input. Percentage of total read counts identified to each library is calculated after unique dual index and inline barcode demultiplexing. Average with perfect normalization is estimated at 1.04% (100/96).

Tunable Coverage for Regions of Interest*

FlexPrep UHT generates high-complexity libraries that have uniform coverage after target enrichment. In combination with Twist Custom Panels, FlexPrep offers tunable coverage of SNPs, k-mers, structural variants, and other genomic areas of interest. Enriched material was downsampled to an average of 75x coverage. Key target enrichment metrics from Picard are reported.

Target Enrichment using a 96-plex

30 ng to 300 ng gDNA Mass Input into Library Preparation

Metrics at 75x Raw Target Coverage

Average +/- Standard Deviation

Selected Bases

Mean Target Coverage

Chimeras

Fold-80 Base Penalty

Covered Bases at 10X

Covered Bases at 20X

Covered Bases at 0X

79.22% +/- 0.54%

32.74 +/- 4.78

1.31% +/- 0.38%

1.416 +/- 0.044

96.06% +/- 0.94%

85.41% +/- 6.36%

0.43% +/- 0.04%

*Method used: 96 libraries were prepared with human genomic DNA (gDNA) (NA12878) using the Twist FlexPrep UHT Library Preparation Kit and FlexPrep Target Enrichment Protocol as a 96-plex with a custom 800 kb panel before sequencing on an Illumina NextSeq 550. Eight library pools of 12 samples each were generated using variable mass input ranging between 30 ng to 300 ng in each library pool. **Standard kits based on workflows which use enzymatic fragmentation followed by ligation and PCR ***All charts, figures, and graphs per Twist internal data September 2024. For research use only. Not for use in any diagnostic or clinical procedures.