The NEBNext Ultra II DNA PCR-free Library Prep Kit contains the enzymes and buffers required to convert a broad range of sheared DNA input amounts into libraries for next-generation sequencing on the Illumina platform, all without an amplification step. The fast, user-friendly workflow requires minimal hands-on time, resulting in excellent library yields and GC coverage without requiring PCR.Advantages
Enjoy flexible input amounts from 250 ng to 1,000 ng of sheared DNA
Get more of what you need with reliably high library yields
Prepare PCR-free libraries with a broad range of sample types
Save time with streamlined workflows, reduced hands-on time, and automation compatibility
Use with the NEBNext Multiplex Oligos for Illumina (Unique Dual Index UMI Adaptors DNA Set 1)or other Illumina-compatible index adaptors with a 3’ single T overhang
A separate version of the NEBNext Ultra II DNA PCR-free Library Prep Kit is available with sample purification beads.
Figure 1: NEBNext Ultra II DNA PCR-free Library Prep Kit generates libraries with higher yieldsA. PCR-free libraries were prepared with NA19240 genomic DNA (Coriell Institute) using NEBNext Ultra II DNA PCR-free and Illumina TruSeq® PCR-free library prep kits and size selected for 350 bp inserts. DNA inputs were 1 µg.B. Libraries of 150-200 bp inserts were prepared using NEBNext Ultra II DNA PCR-free and Roche Sequencing Kapa HyperPrep library prep kits coupled with Covaris shearing without size selection. NEBNext Unique Dual Index UMI Adaptors DNA Set 1, IDT for Illumina (TruSeq DNA UD Indexes) and Kapa Dual-Indexed Adaptors were used for the NEBNext, Illumina, and Kapa kits, respectively, following manufacturers" recommendations.Figure 2: NEBNext Ultra II DNA PCR-free Library Prep provides uniform genome coverage with human DNANEBNext Ultra II DNA PCR-free libraries have even more genome coverage than Illumina DNA PCR-free libraries. Sequencing libraries were prepared according to the manufacturer’s recommendations from 250 ng of NA19240 cell line DNA and sequenced on a single NovaSeq 6000 S4 v1.5 flow cell (PE 2x150 bp). Reads were adaptor trimmed (fastp 0.20.0), aligned to the Telomere-to-Telomere chm13 reference (draft 1, bwa-mem 0.7.17), and duplicate marked (samblaster 0.1.24). 2.8B reads were randomly sampled from each bam (sambamba view -s 0.6.8) Coverage of primary, pass-filter assignments was assessed using mosdepth (v0.3.1, -F 772) and plotted for autosomes or the mitochondrial genome (inset). Expected coverage (num_reads*read length/genome size) is indicated).
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