Bio&SELL 5x QPCR Mix EvaGreen® ROBUST for EvaGreen®-based QPCR is optimized for real-time applications such as quantitative PCR and contains all necessary components for a quantitative PCR except template DNA and primers.
The 5x QPCR Mix EvaGreen® ROBUST is highly specific and provides highly reproducible results with
- low copy numbers
- long templates
- and GC-rich templates
Application:
- the product is mainly used in recurrent deviations with relevant effects on the result
- UNG treatment (dUTP/dTTP in dNTP-Mix)
- extension of nonspecifically bound primers and primer-dimer formation
Devices and Device Settings:
- Compatible with all standard Real-Time cyclers, regardless of the need for Rox
- For all "standard Cyling mode" - "Standard" - and "Fast" - Real-time cyclers
- EvaGreen® is compatible with all commonly used real-time PCR cyclers - simply choose the default setting for SYBR® Green or FAM!
| Product | Art.No. | Quantity |
|---|---|---|
| 5x QPCR Mix EvaGreen® ROBUST | BS76.550.0200 | 200µl = 50 reactions in 20 µl reaction volume |
| BS76.550.1000 | 1.000µl = 250 reactions in 20 µl reaction volume | |
| BS76.550.5000 | 5.000µl = 1,250 reactions in 20 µl reaction volume |
Application
- Detection and quantification of DNA and cDNA targets
- High resolution melt analysis
- Profiling of gene expression
- Microbial detection
- Determination of viral loads
Protocols
Recommended pipetting protocol
| component | volumene | final concentration |
|---|---|---|
| Bio&SELL 5x QPCR Mix ROBUST | 4 µl | 1 x |
| Primer 1 (10 pmol/ μl) |
0,2-0,4 μl |
100 - 200 nM |
| Primer 2 (10 pmol/ μl) |
0,2-0,4 μl |
100 - 200 nM |
| Template-DNA | 1-5 µl | 0,002 – 2 ng/μl |
| Water | fill up to 20 µl |
Recommended QPCR protocol: Initial incubation for 12 minutes at 95 ° C
| component | temperature | time | cycles |
|---|---|---|---|
| Initiale Incubation | 95°C | 12 min | 1 x |
| Denaturation | 95°C | 12-15 sec. | 40 x |
| Annealing | 59 - 65°C | *19-30 sec. | |
| Elongation | 72°C | *19-30 sec. | |
| * 19 sec sufficient for 150bp templates | |||
EvaGreen® is a DNA-binding dye with many advantages over SYBR® Green I in quantitative PCR. Although on a similar spectrum, EvaGreen® has the following important advantages over SYBR® Green I :
EvaGreen® has a lower PCR inhibition; is an extremely stable dye and is neither mutagenic nor cytotoxic. EvaGreen® is compatible with all common real-time PCR cyclers - simply select the default setting for SYBR® Green or FAM!
With internal reference dye ROX. ROX is a passive internal reference dye that is used for normalization of the fluorescent reporter signal during QPCR. The final concentration of ROX dye in the reaction mixture is 500 nM.
*EvaGreen® is a registered trademark and licensed for sale by BIOTIUM, Inc., Hayward, CA, USA.
This product is provided under an agreement with BIOTIUM, INC., and the manufacture, use, sale or import of this product is subject to one or more of pending patent applications owned by BIOTIUM and ALLELOGIC. The purchase of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product in research conducted by the buyer, where such research does not include testing, analysis or screening services for any third party in return for compensation on a per test basis. The buyer cannot sell or otherwise transfer (a) this product (b) its components or (c) materials made using this product or its components to a third party or otherwise use this product or its components or materials made using this product or its components for Commercial Purposes. Commercial Purposes means any activity by a party for consideration and may include, but is not limited to: (1) use of the product or its components in manufacturing; (2) use of the product or its components to provide a service, information, or data; (3) use of the product or its components for therapeutic, diagnostic or prophylactic purposes; or (4) resale of the product or its components, whether or not such product or its components are resold for use in research. For information on purchasing a license to this product for purposes other than research, contact BIOTIUM, INC., Business Development, 3423 Investment Blvd, Suite 8, Hayward, CA 94545 Tel: 510-265-1027, Fax: 510-265-1352.
SYBR® is a registered trademark of Molecular Probes, Inc.