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Invitrogen™ SYBR GreenER™ qPCR SuperMix Universal
Description
Includes
2X SYBR GreenER qPCR SuperMix and a separate tube of ROX reference dye
SYBR™ GreenER™ qPCR SuperMixes Universal incorporate the latest high-performance technology to provide the most reliable gene expression data on a variety of platforms, including ABI 7500, Corbett Rotor-Gene™, MJ Chromo4™ and Opticon™, and Stratagene instruments. The SYBR™ GreenER™ system is specially formulated to offer the best sensitivity, specificity, and reproducibility with. A novel dsDNA-binding dye exhibits a brighter signal for increased sensitivity and less PCR inhibition than original SYBR™ Green I, while using the same instrument filters and settings. An optimized buffer system improves sensitivity and provides excellent long-term stability. Uracil DNA glycosylase reduces carryover contamination in qPCR.
Eight cDNA targets and eight genomic DNA targets were amplified from 1ng of cDNA and 6pg of gDNA, respectively, on the ABI PRISM™ 7900HT Sequence Detection System using the SYBR™ GreenER™ qPCR SuperMix for ABI PRISM™ instruments. For all 16 targets, detection was positive in the presence of template but negative for all no-template controls.
- The SYBR™ GreenER™ qPCR SuperMix kits include a new dsDNA-binding dye that produces a brighter signal and significantly less PCR inhibition, for improved qPCR performance over a broad dynamic range
Consistent Specificity and Low-Copy Detection:
- The SYBR™ GreenER™ qPCR reagent system minimizes the formation of nonspecific products, including primer-dimers, for greater accuracy across a wide range of targets
- The reliability of the SYBR™ GreenER™ qPCR reagent system also translates to greater sensitivity
- With SYBR™ GreenER™ qPCR SuperMix, you can consistently detect fewer than 10 copies of a target in genomic DNA
Specifications
Specifications
| Concentration | 2X |
| Content And Storage | Contains SYBR™ GreenER™ qPCR SuperMix Universal (12.5 mL). SYBR™ GreenER™ qPCR SuperMix Universal is a 2X SuperMix with ROX Reference dye provided in a separate tube (500 μL). Sufficient reagents are provided for 500 reactions based on a 50 μL reaction size. Store at +4°C upon receipt. |
| Detection Method | SYBR |
| Format | Tube |
| GC-Rich PCR Performance | High |
| PCR Method | qPCR |
| Reaction Speed | Standard |
| For Use With (Equipment) | 7500 System, Stratagene Mx4000, Stratagene Mx3000P, Stratagene Mx3005P, MJ Chromo4, MJ Opticon |
| Product Line | Platinum, SYBR GreenER |
| Product Type | Real Time PCR SYBR Master Mix |
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Frequently Asked Questions (FAQs)
Sensitivity can actually be equivalent when using TaqMan chemistry and SYBR Green reagent chemistry. It might seem that a TaqMan assay with fluorescent signal generated by a sequence-specific probe would always be more sensitive than a SYBR Green reagent assay, but a poorly designed TaqMan assay could theoretically be less specific than a well-designed SYBR Green reagent assay. However, the potential for detection of primer dimers and non-specific products using SYBR Green chemistry is more likely to result in loss of sensitivity when attempting to quantitate lower copy numbers.
For more information on Real-Time PCR chemistries, please refer to the following Application Notes, which you can find on our website through Technical Resources, or by entering the titles in the main Search field: “Real-Time PCR Vs. Traditional PCR”, “Essentials of Real Time PCR”, and “Selection of Reagents for Real-Time PCR”.
The TaqMan MGB probes contain the following features:
1) A fluorescent reporter at the 5' end
2) A nonfluorescent quencher at the 3' end. Because the quencher does not fluoresce, the real-time instruments can measure the reporter dye contributions more precisely.
3) A minor groove binder at the 3' end. The minor groove binder increases the melting temperature (Tm) of probes, allowing the use of shorter probes.
In general, the TaqMan MGB probes exhibit great differences in Tm values between matched and mismatched probes, which provides more accurate allelic discrimination and makes for a more sensitive real-time assay. Mismatches between a probe and allele, or target, reduce the efficiency of probe hybridization in a measurable way, which is especially important in SNP Genotyping assays. Furthermore, AmpliTaq Gold DNA polymerase is more likely to displace the mismatched probe rather than cleave it to release reporter dye. More information about TaqMan MGB probes can be found in the User Bulletin entitled "Primer Express Version 1.5 and TaqMan MGB Probes for Allelic Discrimination." You can find a copy on our website by entering this title in the main search field.
Refer to the product manual for your instrument and software for specifics, but in general you will want to change the Quencher value to None.
It is generally not possible to use DNA as a standard for absolute quantitation of RNA because there is no control for the efficiency of the reverse transcription step. Therefore, in-vitro transcribed RNA is commonly used to prepare standards for the absolute quantitation of RNA targets. This would involve the cloning of the target of interest into an in-vitro transcription plasmid, performing in-vitro transcription, then purifying the resulting cRNA so that the DNA plasmid cannot serve as a PCR template. Concentration is measured by A260 and converted to the number of copies using the molecular weight of the RNA.
Relative quantitation of gene expression methods require less up-front preparation and provide a fold-change value instead of an absolute quantity result. For many researchers, absolute quantities are not a necessary parameter to measure, and therefore relative quantitation is a much more attractive approach to studying gene expression via real-time PCR. For more information on relative quantitation of gene expression, please refer to our Technical Reference Library in the Technical Resources section of our website.
Safety and Handling
For Research Use Only. Not for use in diagnostic procedures.
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