Hieff NGS™ OnePot Pro DNA Library Prep Kit V3 is a next-generation enzymatic library preparation kit compatible with both Illumina and MGI high-throughput sequencing platforms. Compared to traditional library construction methods, this kit replaces time-consuming and instrument-dependent sonication with a high-quality, low-bias fragmentation enzyme. It uniquely integrates the fragmentation, end repair, and dA-tailing steps into a single reaction, significantly reducing hands-on time, cost, and workflow complexity. The ligation module features pre-mixed enzyme and buffer for streamlined operation, making the kit highly suitable for automation.

Building upon the previous version, this updated formulation further reduces GC bias, enhances end-repair and dA-tailing efficiency, and improves reagent stability. It also incorporates a newly optimized ligase for superior adapter ligation performance. The kit is fully compatible with standard Illumina or MGI adapters and primers. Ideal for whole-genome sequencing of animals, plants, microbes, and challenging clinical samples.

Features

Wide input range: Compatible with 1 ng–1 µg of genomic DNA, full-length cDNA, or other double-stranded DNA

High-quality fragmentation enzyme: Enables random dsDNA cleavage with minimal sequence bias, integrating fragmentation, end repair, and dA-tailing in a single step

High-fidelity, high-efficiency PCR enzyme: Maximizes library yield and complexity while preserving sequence accuracy

FFPE-compatible: Delivers robust performance with degraded or formalin-fixed DNA samples

Rigorous quality control: Each production batch undergoes stringent performance validation and stability testing

Specifications

Components

[Note]: The kit components are compatible with both Illumina &MGI sequencing platform, if the complete adapter was used, Hieff NGS™ Primer Mix (Yeasen Cat#12190 or Cat#12191) in needed.

Application

DNA library Preparation; Whole genome sequencing(WGS); whole exome or other targeted capture sequencing; amplicon sequencing; immunoprecipitation sequencing; metagenomic sequencing;

Figures

1. Workflow

Figure 1. The workflow of OnePot Pro DNA Library Prep Kit

2. Flexible and High-Efficiency Fragmentation

Figure 2. Fragment size distribution of 250 ng human gDNA using the 12194 fragmentation module.

Calf gDNA (250 ng) was fragmented at 30°C ~35°C for 5 min, 10, 15, 20, 30. The distribution of recovered insert sizes is shown, demonstrating stable and uniform fragmentation across the 5–30 minute range.

3. Broad Species Compatibility

Figure 3. Library uniformity testing across different species using 12194 under identical fragmentation conditions.igure 3. Li

Different DNA samples were fragmented with 12194 under identical conditions. The libraries showed consistent sizes (~450 bp / ~320 bp) with good uniformity across species.

Figure 4. Comparison of soft-clipping rates (%) and false chimeric read rates (%) across human gDNA, ctDNA, and FFPE DNA samples. The results show that 12194 exhibits lower or comparable soft-clipping rates (%) and false chimeric rates(%).

5. Strong Stability

Figure 5. Library yield stability after storage at 4°C and 25°C for 4 weeks. No decrease in library yield was observed after 4 weeks of storage at 4°C and 25°C, highlighting the kit’s suitability for automation.