NadPrep Methyl Library Preparation Kit v2Introduction NadPrep Methyl Library Preparation Kit v2 is designed for preparation of high-quality methylation sequencing (Methyl-Seq) libraries from double-stranded DNA (dsDNA) on both Illumina and MGI platforms. This kit is suitable for preparing Methyl-Seq libraries from 1-500 ng DNA, supporting whole-genome bisulfite sequencing (WGBS) as well as hybrid capture-based targeted sequencing with effective enrichment of methylated libraries. It is compatible with various types of DNA samples, including gDNA, cfDNA, and FFPE DNA, as well as a wide range of commercially available conversion solutions. Compared to the v1 version, the upgraded kit shortens experimental time, supports lower sample input amounts, and significantly improves library yield and data utilization efficiency, offering users a more efficient, flexible, and reliable methylation sequencing solution. • Supports input amounts ranging from 1-500 ng across different DNA sample types, enabling better sample utilization. • Compatible with various conversion methods and sequencing platforms, offering flexibility for diverse applications. • Enables WGBS library preparation in as fast as 4.5 hr with bisulfite conversion. • Significantly improves effective sequencing depth and data utilization. Library Amplification Post-amplification Cleanup

Efficient Library Yield Figure 1. NadPrep Methyl Library Preparation Kit v2 achieves stable and efficient library yield across various applications. A. gDNA samples of varying input amounts processed with NadPrep Methyl Library Preparation Module v1 and v2 after BS conversion; B. gDNA samples of varying input amounts with EM; C. Library preparation for gDNA, FFPE DNA, and cfDNA with BS; D. Simulated samples with varying methylation levels with BS. Note: The samples in figure 1.D were simulated DNA by using human 100% Methylated DNA standard (Zymo, D5014-2) positive control and 0% Methylated DNA...

Improved Data Utilization Average depth (rmdup/x) Normalized coverage Target covered (%) NadPrep v1-100 ng NadPrep v2-10 ng NadPrep v2-50 ng Figure 3. Capture performance of NadPrep Methyl Library Preparation Kits v1 and v2. A. Mappability & On-target rate; B. Target covered; C. Average sequencing depth after deduplication; D. GC bias. Pre-libraries were prepared from gDNA samples with various input amounts using NadPrep Methyl Library Preparation Module v1 & v2 coupled with NadPrep Methyl Stubby Adapter (UDI) Module (with 10 nt Index) and treated by BS. Hybrid capture was performed with NadPrep...

Flexible Compatibility with Ultra-Sensitive μCaler Hybrid Capture System Target covered (%) Average depth (rmdup/x) Figure 5. NadPrep Methyl Library Preparation Kit v2 demonstrates excellent capture performance with low-input gDNA samples. A. Library yield; B. Mappability & On-target rate; C. Target covered; D. Average sequencing depth after deduplication. Pre-libraries were prepared using NadPrep Methyl Library Preparation Module v2 coupled with NadPrep Methyl Stubby Adapter (UDI) Module (with 10 nt Index) and treated by EM. Hybrid capture was performed with μCaler Hybrid Capture Reagents v2...