NGS Library Quantitation

Accurate quantitation of next-generation sequencing (NGS) libraries is essential for optimizing data yield and quality from each sequencing run. Precise quantitation ensures that the appropriate amount of library is loaded onto the NGS platform, facilitating reliable predictions of sequencing output. Quantitative PCR (qPCR) is recognized as the most accurate and effective method for library quantitation, offering significantly higher consistency and reproducibility compared to alternative techniques. Amplification based methods, such as qPCR, target adaptor containing sequences, providing a more accurate estimate of the concentration of library molecules available for sequencing. NEBNext Library Quant kits are optimized with platform-specific reagents to ensure efficient amplification of Illumina^® or Ultima Genomics^® NGS libraries. The streamlined protocol minimizes pipetting steps and uses a single extension time for all libraries, enhancing efficiency while generating reproducible, high-quality results.

The NEBNext^® Library Quant Kit for Illumina^® is designed for precise qPCR-based quantitation of NGS libraries prepared for Illumina sequencing platforms. This kit includes primers targeting the P5 and P7 Illumina adaptor sequences and pre-diluted standards for reliable quantitation of DNA libraries ranging from 150 to 1,000 bp. It offers flexibility with the option to use either four or six standards. ROX is included in the kit for use with qPCR instruments that require a reference dye for normalization.

The NEBNext^® Library Quant Kit for Ultima Genomics^® delivers precise and reliable qPCR-based quantitation of NGS libraries prepared using the UG 100 Solaris™ Free workflow. The kit features forward and reverse primers targeting the Ultima adaptor sequences, and includes five high-quality, pre-diluted DNA standards for reliable quantitation of diluted DNA libraries. Additionally, it incorporates the Luna^® Universal qPCR Master Mix with a blue, non-interfering, visible tracking dye, which can also serve as a passive reference dye, eliminating the need for additional ROX in real-time instruments requiring ROX normalization.

For more information about the importance of quantitation, read "The Quantitation Question: How does accurate library quantitation influence sequencing?".