sparQ DNA Frag & Library Prep Kit

Robust enzymatic fragmentation & consistent results

Features & Benefits

  • Simple 2 step workflow employs a unique enzyme mix safeguarding samples from over fragmentation
  • Tunable and reproducible fragmentation profiles
  • Flexible generation of high quality libraries from 1 ng - 1 µg of input DNA
  • PCR-free workflows enabled from 100 ng
  • Minimized bias across challenging regions for improved sequencing results

 

sparQ DNA Frag & Library Prep Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention or treatment of a disease.

Description

The sparQ DNA Frag & Library Prep Kit optimizes the integration of enzymatic fragmentation into a two-step protocol for the streamlined construction of libraries for sequencing on Illumina® NGS platforms. A single-tube enzyme mix facilitates the combination of fragmentation and DNA polishing minimizing DNA over fragmentation while greatly simplifying library. The streamlined workflow can be completed in under 3 hours with minimal hands-on time and can accommodate DNA input amounts from 1 ng to 1000 ng. DNA fragmentation and DNA polishing enzymes work together generating fragment sizes that are tunable and reproducible based on reaction time. Double-stranded DNA molecules are fragmented followed by DNA polishing reactions where 5'-phosphorylated and 3'-dA-tailed DNA fragments suitable for direct ligation of sequencing adapters are generated without the need for an intervening cleanup step. The HiFi PCR Master Mix and Primer Mix allow unbiased amplification of fragments with appropriate adapters ligated to both ends. PCR-Free workflows are enabled from 100 ng of starting material.

 

Streamlined  workflow

sparQ DNA Frag& Library Prep workflow


sparQ DNA Frag & Library Prep workflow

The streamlined workflow utilizes a proprietary enzyme mix that integrates tunable and reproducible fragmentation with DNA polishing simplifying library construction. The same single reaction tube is used to proceed to adapter ligation and cleanup, minimizing sample transfer steps. A second tube is used for workflows requiring PCR amplification, and a final tube receives the sequencing-ready library.


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  • Enzyme mix integrates fragmentation and DNA polishing reactions for a streamlined workflow
  • Complete workflow in under 3 hours with minimal hands on time


Tunable fragmentation

Fragmentation tuning guide


Fragmentation tuning guide

Simply select the desired fragment size and input DNA amount. If input DNA falls between values displayed on the graph, an estimate can be used for optimizing fragmentation times.


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  • Adjusting fragmentation time generates selective DNA sizes
  • Produce more usable library fragments in the target range with sharper size distribution curves

Fragmentation time course

Fragmentation time course


Fragmentation time course

sparQ DNA Frag & Library Prep Kit is tunable to the desired fragment size. 100 ng Human gDNA was subjected to fragmentation with a series of incubation time points (3 – 18 min). After fragmentation, DNA samples were purified and then visualized using Agilent High Sensitivity DNA Kit.


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  • Select fragment size and input DNA amount to determine reaction times
  • Optimize fragmentation tuning for your sample


Minimize bias and maintain genome diversity

Genome coverage analysis


Genome coverage analysis

Library prepared using sparQ DNA Frag & Library Prep Kit resulted in uniform coverage across the wide range of GC-content. Libraries were prepared using different DNA fragmentation and library preparation kits with 1 ng or 100 ng of microbial gDNA followed by sequencing on Illumina MiSeq. 2 million reads from each tested library were down-sampled and analyzed. Coverage uniformity against GC-content bias resulting from different DNA fragmentation and library preparation kits were compared by plotting normalized coverage for a wide GC-content.


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  • Low fragmentation and amplification bias enables better coverage uniformity
  • Consistent coverage independent of input DNA amounts
  • PCR-Free sequencing data correlates well with mechanical shearing results


Quality sequencing metrics

Quality sequencing metrics and improved workflow economics


Quality sequencing metrics and improved workflow economics

sparQ DNA Frag & Library Prep Kit generates high quality DNA libraries with minimal duplication artifacts. Libraries were prepared with 1 ng and 100 ng of microbial genomic DNA, amplified for 12 and 6 cycles respectively, and subsequently sequenced on Illumina MiSeq. Each library was down-sampled to 2 million reads (150 bp paired-end reads) and aligned to a reference genome with only unique alignments reported.


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  • Industry leading sequencing results
  • High mapped reads with minimal duplication rates
  • Ensured return on sequencing investment


sparQ Adapter Barcode Sets

sparQ Adapter Barcode Sets A and B are single index barcoded oligonucleotide adapters designed for use with the Quantabio sparQ DNA Library Prep Kits in the construction of libraries for Illumina NGS platforms. Each sparQ Adapter Barcode Set contains 12 single index barcodes for 96 rxns.

sparQ Adapter Barcode Set A
Barcode Number      Barcode Sequence
2 CGATGT
4 TGACCA
5 ACAGTG
6 GCCAAT
7 CAGATC
12 CTTGTA
13 AGTCAA
14 AGTTCC
15 ATGTCA
16 CCGTCC
18 GTCCGC
19 GTGAAA
sparQ Adapter Barcode Set B
Barcode Number  Barcode Sequence
1 ATCACG
3 TTAGGC
8 ACTTGA
9 GATCAG
10 TAGCTT
11 GGCTAC
20 GTGGCC
21 GTTTCG
22 CGTACG
23 GAGTGG
25 ACTGAT
27 ATTCCT

Performance Data

Fragmentation time course


Fragmentation time course

sparQ DNA Frag & Library Prep Kit is tunable to the desired fragment size. 100 ng Human gDNA was subjected to fragmentation with a series of incubation time points (3 – 18 min). After fragmentation, DNA samples were purified and then visualized using Agilent High Sensitivity DNA Kit.


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Streamlined workflow


Streamlined workflow

The streamlined workflow utilizes a proprietary enzyme mix that integrates tunable and reproducible fragmentation with DNA polishing simplifying library construction. The same single reaction tube is used to proceed to adapter ligation and cleanup, minimizing sample transfer steps. A second tube is used
for workflows requiring PCR amplification, and a final tube receives the sequencing-ready library.


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Fragmentation tuning guide


Fragmentation tuning guide

Simply select the desired fragment size and input DNA amount. If input DNA falls between values displayed on the graph, an estimate can be used for optimizing fragmentation times.


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Superior quality libraries & yields


Yield comparison

sparQ DNA Frag & Library Prep Kit shows significantly higher NGS library preparation efficiency. Libraries with 300 bp average DNA fragments from 100 ng of gDNA Coriell NA12878 were prepared using sparQ DNA Frag & Library Prep Kit and a commercial kit. Manufactures’ manuals were carefully followed. Amplified libraries (5 cycles of amplification) were quantified by Qubit fluorometric quantitation method


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Genome coverage analysis


Genome coverage analysis

Library prepared using sparQ DNA Frag & Library Prep Kit resulted in uniform coverage across the wide range of GC-content. Libraries were prepared using different DNA fragmentation and library preparation kits with 1 ng or 100 ng of microbial gDNA followed by sequencing on Illumina MiSeq. 2 million reads from each tested library were down-sampled and analyzed. Coverage uniformity against GC-content bias resulting from different DNA fragmentation and library preparation kits were compared by plotting normalized coverage for a wide GC-content.

Libraries prepared using PCR-free workflow of sparQ DNA Frag & Library Prep Kit with 50 ng of microbial genomic DNA shows similar high performance as a typical amplified library prepared by Covaris mechanical shearing method.


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Quality Sequencing metrics and improved workflow economics


Quality Sequencing metrics and improved workflow economics

sparQ DNA Frag & Library Prep Kit generates high quality DNA libraries with minimal duplication artifacts. Libraries were prepared with 1 ng and 100 ng of microbial genomic DNA, amplified for 12 and 6 cycles respectively, and subsequently sequenced on Illumina MiSeq. Each library was down-sampled to 2 million reads (150 bp paired-end reads) and aligned to a reference genome with only unique alignments reported.


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What Customers Say
The sparQ kit safeguards samples from over fragmentation and routinely provides more consistent fragment size. Once optimized, I had peace of mind knowing my... Read More The sparQ kit safeguards samples from over fragmentation and routinely provides more consistent fragment size. Once optimized, I had peace of mind knowing my samples were safe when I got busy in the lab.”
Scientist, Laboratory Manager
Agricultural genomics institute

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