Library Quantification Kit
Library Quantification Kit
Brand: Takara Bio.
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Library Quantification Kit
NGS library quantification kit
The Library Quantification Kit is used for rapid library quantification prior to Illumina next-generation sequencing. The kit contains all of the components needed to determine the amount of library DNA (genomic, cDNA, etc.) in a sample using qPCR. Quantification is accomplished by comparison of the threshold cycle (Ct) of the diluted sample with a standard curve derived from four calibrated DNA standards.
The DNA Standards for Library Quantification includes four DNA standards (0.01–10 pM) that are used for qPCR-based library quantification prior to Illumina next-generation sequencing. These DNA standards contain specific concentrations of a DNA fragment (538 bp) and are designed specifically for use with the Library Quantification Kit (Cat No. 638324). When used as templates for qPCR with the Library Quantification Kit, the standards will generate 447-bp DNA fragments that can be used to produce a standard curve for determining the concentration of DNA in an Illumina library. The standards are not suitable for use with other qPCR techniques, as they may lead to quantification errors or poor results.
Overview
- Highly sensitive quantification of Illumina sequencing libraries—the qPCR-based assay can measure libraries at low concentrations
- Allows specific quantification of only adapter-bound DNA molecules
- Reduces variability in cluster density to improve sequencing results—use prior to sequencing to ensure optimal loading concentration
- Complete kit—includes qPCR primers and reagents, and DNA standards for absolute quantification using a standard curve
Applications
- Illumina NGS library quantification by qPCR
Lot-to-lot variation for DNA Standards
Lot-to-lot variation for DNA Standards. Quantification of three lots of pre-diluted DNA Standards (0.01, 0.1, 1, and 10 pM) was performed by real-time PCR using Terra qPCR Direct TB Green Premix following the protocol in the Library Quantification Kit user manual. All reactions were performed in triplicate, and the resulting Ct (SDM) values for the triplicates were averaged. The variation between lots was <5%.
A typical RNA-seq workflow
A typical RNA-seq workflow. Generating sufficient amounts of library DNA and loading an appropriate amount on the flow cell is essential for obtaining high-quality NGS data. The Library Quantification Kit, a qPCR-based quantification method that targets Illumina’s adapter sequences, can be used to measure library concentration and ensure optimal loading concentration.
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Lot-to-lot variation for DNA Standards
Lot-to-lot variation for DNA Standards. Quantification of three lots of pre-diluted DNA Standards (0.01, 0.1, 1, and 10 pM) was performed by real-time PCR using Terra qPCR Direct TB Green Premix following the protocol in the Library Quantification Kit user manual. All reactions were performed in triplicate, and the resulting Ct (SDM) values for the triplicates were averaged. The variation between lots was <5%.
A typical RNA-seq workflow
A typical RNA-seq workflow. Generating sufficient amounts of library DNA and loading an appropriate amount on the flow cell is essential for obtaining high-quality NGS data. The Library Quantification Kit, a qPCR-based quantification method that targets Illumina’s adapter sequences, can be used to measure library concentration and ensure optimal loading concentration.
