Library quantifcation master mix for Illumina®
Accurate quantitation of the number of molecules in a library is the most critical step in the preparation steps for next-generation sequencing (NGS) to obtain high-quality reading data. Indeed, in order to obtain reliable and quality reading data, it is necessary to use a sufficient quantity of DNA, and in particular for Illumina® sequencing. The use of insufficient DNA will result in low sequencing efficiency and DNA overabundance will increase cluster density and result in poor quality data.
Quantitative PCR is considered to be the most accurate and efficient method of quantification of NGS library.
The library quantitation kits for Illumina® sequencing contain all the components bellow:
- DNA polymerase
- Primers to amplify only the molecules that contain the two sequences of adapters
- SYBR green or EvaGreen
- A ROX solution to add depending on the device used. If the ROX solution is separate from the mix, it is a universal kit that can be used with all instruments. If ROX is contain in the master mix, it is necessary to use the appropriate kits with your device.
|Without ROX||Bio-Rad CFX96, CFX384, iCycler iQ, iQ5, MyiQ, MiniOpticon, Opticon®, Opticon 2, Chromo4;|
Cepheid SmartCycler®; Eppendorf Mastercycler® ep realplex, realplex 2 s;
Illumina Eco qPCR;
Qiagen/Corbett Rotor-Gene® Q, Rotor-Gene® 3000, Rotor-Gene® 6000;
Roche Applied Science LightCyclerTM 480;
Thermo Scientific PikoReal Cycler.
|Low ROX||Applied Biosystems 7500, 7500 Fast, ViiA7; Stratagene MX4000,|
|High ROX||Applied Biosystems 5700, 7000, 7300, 7700, 7900, 7900HT, 7900HT Fast;|
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