HighPrep PCR PB is bead based reagent designed for efficient removal of DNA molecules less 5 kb and library size selection during SMRTbell® library preparation. It also cleans amplicon template for SMRT library preparation by the removal of salts, PCR buffers, excess primers, primer-dimers, dNTPs and non-specific products of PCR. Furthermore, it can be used for purifying a low purity DNA sample prior to library preparation process. Overall, HighPrep PCR PB is an enhanced direct replacement of AMPure PB.
Summary: Benefits of Using HighPrep PCR PB
  • Cost effective when compared with competitors in the market
  • Alternative to AMPure PB-no protocol change
  • Amenable to automation
  • High DNA recovery if quality input DNA is used
  • Removal of DNA fragments less than 5 kb
  • Efficient DNA and amplicon purification
  • Concentration of DNA from highly diluted samples
Library Preparation
Materials
  • Whole blood sample
  • HMW DNA extraction kit: HighPrep HMW DNA Kit Cat#
  • Quality gDNA with A260/A80=1.8 and A260/A230= 2.0
  • Sheared gDNA modal size between 15 -18 kb using the Diagenode Megaruptor 3 system
  • QC of input genomic DNA and final SMRTbell library on the Agilent Femto Pulse system
  • SMRTbell prep kit 3.0 P/N 102-182-700
  • Sequencing Plate: Revio Sequencing plate P/N 102-587-400
  • HighPrep PCR PB Cat# PB-60005
  • AMPure PB P/N 100-265-900
  • Qubit 1x dsDNA HS assay
Method
High molecular weight DNA was extracted from human whole blood using HighPrep HMW DNA Kit. DNA quality was assessed on Agilent Femtopulse system and then shreaded to 15-18 kb using Diagenode Megaruptor 3 system.
This was followed by removal of small fragments (<5 kb) using either HighPrep PCR PB or AMPure PB.
Sheared DNA was used in SMRT library preparation where HighPrep PCR PB and AMPure PB were used in DNA cleanup and size selection.
Results
1. DNA Extraction with HighPrep HMW DNA Kit

Figure 1:

2. Shreaded DNA Sample Profile

Figure 2:

3. HiFi Sequencing QC Metrics

Table 1: GC Bias Analysis

Table 2: HiFi Read Statistics

Conclusions
Our study demonstrates that HighPrep PCR PB is highly suitable for integration into PacBio SMRT sequencing workflows, consistently delivering expected results. Specifically, HighPrep PCR PB outperformed AMPure PB in terms of N50 and mean read length, showcasing its superior performance metrics. This makes HighPrep PCR PB not only a viable but also an enhanced and cost-effective alternative to AMPure PB, ensuring high-quality sequencing outcomes at a reduced cost.
Request a sample of HighPrep PCR PB to experience its benefits firsthand and explore its compatibility with your sequencing projects at: https://www.magbiogenomics.com/request-a-sample