Accurate library quantification for next gen sequencing
PerfeCta NGS Quantifications kits use real-time PCR with prediluted standards for the convenient and accurate quantification of NGS libraries for the Ion Torrent or Illumina NGS platforms.
Features and Benefits
- Accurate and sensitive method for NGS library quantification
- Stabilized, prediluted standards for convenient use
- Consistency across a broad range of samples
- Sensitivity of detection to quantify low concentration libraries
- Quantification for either Illumina or Ion Torrent
Download the PerfeCTa NGS Quantification Brochure
Accurate quantification of the number of amplifiable library molecules is the most critical step in the NGS workflow in obtaining high quality read data with next-generation sequencing technologies. The PerfeCta NGS Library Quantification Kit uses real-time PCR to specifically quantify library molecules that possess the appropriate adapter tag at each end. These are the suitable template molecules for either emulsion PCR used for the Ion Torrent platform or Bridge PCR used for Illumina NGS platforms.PerfeCta NGS Quantification kits simplify the library quantification process by providing stabilized, pre-diluted standards, pre-qualified primer sets, and an optimized dilution buffer for your NGS library samples. This minimizes pipetting errors and ensures reproducible and precise qPCR results, even with dilute samples. The robust qPCR performance of PerfeCta SYBR Green SuperMix provides accurate quantification of NGS libraries with varying fragment sizes or GC content. Kits are available to support all major qPCR instrument platforms.A common problem with some NGS library quantification protocols is the use of DNA standards that are too concentrated and generate qPCR data that are outside of the linear dynamic range for many qPCR instruments. Improper baseline settings result in compressions between the highest concentrated DNA standards that in turn give rise to inflated PCR efficiencies and inaccurate library quantification results. The NGS DNA standards supplied with the PerfeCta NGS Library quantification kits have been carefully selected to avoid these artifacts and produce NGS library standard curves with exceptionally high linear regression correlation coefficients.
The DNA standard for Illumina NGS platforms generates a 426-bp amplicon (48.8% GC). Primer sequences correspond to the “P5″ and “”P7” primer sequences for Illumina sequencing libraries:
Illumina forward: 5’-AATGATACGGCGACCACCGA-3’
Illumina reverse: 5’-CAAGCAGAAGACGGCATACGA-3’
Real-Time PCR Instrument Compatibility
- Roche LightCycler™ 480
- Qiagen/Corbett Rotor-Gene® Q
- Qiagen/Corbett Rotor-Gene® 6000
- Qiagen/Corbett Rotor-Gene® 3000
- Eppendorf Mastercycler® ep realplex
- Eppendorf Mastercycler® ep realplex2 s
- Cepheid SmartCycler®
- Bio-Rad CFX96™
- Bio-Rad CFX384™
- Bio-Rad/MJ Opticon®
- Bio-Rad/MJ Opticon 2
- Bio-Rad/MJ Chromo4™
- Bio-Rad MiniOpticon™
- Illumina Eco qPCR
- Thermo Scientific PikoReal Cycler
- BioRad iCycler iQ™
- BioRad MyiQ™
- BioRad iQ™5
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Kit components are stable for 2 years when stored in a constant temperature freezer at -20°C and protected from light*. For convenience, they may be stored unfrozen at +2 to +8°C for up to 6 months. Repeated freezing and thawing will not affect product performance.
*Prolonged exposure of PerfeCTa SYBR Green SuperMix to direct light will photo-bleach the SYBR Green I dye and destroy qPCR product detection.
Precautions and Disclaimer
For research use only. Not intended for any animal or human therapeutic or diagnostic use.
Please consult the Material Safety Data Sheet for information regarding hazards and safe handling practices.
|Cat# 95154-500||500 x 20 µl reactions|