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| Product Name | Universal Blue qPCR SYBR Green Master Mix (with ROX) |
| Catalog No. | RESK-0003 |
| Description | This 2× ready-to-use premix for real-time quantitative PCR combines high sensitivity with high specificity, and its blue color provides a convenient visual indicator for sample loading. The core component, an antibody-mediated hot-start Taq DNA polymerase, effectively suppresses non-specific amplification caused by primer annealing during reaction setup. The formulation also contains factors that enhance PCR amplification efficiency and balance the amplification of genes with different GC contents (30-70%), delivering reliable linearity across a wide quantitative range. A specially formulated ROX passive reference dye is premixed into the reagent, making it compatible with all qPCR instruments without any need to adjust ROX concentration for different platforms — simply add primers and template to the reaction mix and start amplification. |
| Intended Use | Dye-based real-time quantitative PCR (qPCR) for gene expression analysis and nucleic acid quantification, universally compatible with all qPCR instrument platforms. |
| Principle / Technology | Antibody-mediated hot-start Taq DNA polymerase with SYBR Green I dye chemistry; a premixed universal ROX passive reference dye eliminates platform-specific ROX adjustment, while a blue tracking dye allows visual confirmation of sample loading. |
| Detection Method | Fluorescent quantitative PCR (SYBR Green dye-based) |
| Sample Type | cDNA and DNA templates (e.g., plasmid DNA) |
| Performance Range / Specifications | Effectively detects template amounts across 7 orders of magnitude (10^0-10^6 plasmid copies) with high amplification efficiency and excellent linearity. Precisely distinguishes 2-fold differences in template concentration. Outstanding reproducibility: amplification curves from 90 replicates highly overlap, with a Ct standard deviation below 0.2. Reaction mixes prepared at room temperature can be stored at 4°C for 24 hours with no change in detection results. Compatible with standard and fast programs, completing quantification in as little as 46 minutes. Earlier Ct values than a leading competing brand across different instrument platforms. |
| Sensitivity / LOD | High sensitivity, capable of detecting down to a single-copy gene. |
| Specificity | Antibody-mediated hot-start Taq DNA polymerase effectively suppresses non-specific amplification during reaction setup; validated across 200-bp amplicons with 30-70% GC content, ensuring a high specific detection rate. |
| Reaction Conditions / Protocol | A two-step protocol is recommended for routine use; switch to a three-step protocol or extend the extension time when amplification efficiency is low and Ct values are high. Compatible with both standard and fast cycling programs, with quantification completed in as little as 46 min. |
| Components / Formulation | 2× blue qPCR premix containing antibody-mediated hot-start Taq DNA polymerase, SYBR Green I fluorescent dye, universal ROX passive reference dye, and a blue tracking dye. Only primers and template need to be added by the user. |
| Storage Conditions | Store at -20°C, protected from light. Avoid repeated freeze-thaw cycles. The product contains the fluorescent dye SYBR Green I; avoid strong light exposure during storage and reaction setup. |
| Shelf Life | 18 months |
| Package Specifications | 1 mL; 5 × 1 mL; 50 × 1 mL; 100 × 1 mL |
| Product Form | 2× premix |
| Concentration | 2× |
| Shipping Conditions | Shipped with ice packs. |
| Expiration Date / Stability | Stable for 18 months at -20°C protected from light. Prepared reaction mixes remain stable at 4°C for 24 hours with unchanged detection results. A pale-yellow precipitate may form after frozen storage due to reduced stabilizer solubility; this does not affect performance — mix by inversion and gentle pipetting until the precipitate dissolves. |
| Compatibility | Universal for all qPCR instrument platforms thanks to the premixed special ROX reference dye; no ROX concentration adjustment is required. Compatible with standard and fast qPCR programs. Recommended amplicon length 80-300 bp. |
| Application Notes / Precautions | A two-step protocol is commonly used; to improve specificity, raise the annealing temperature; for low efficiency or high Ct values, switch to a three-step protocol or extend the extension time. Valid qPCR results require: amplification efficiency of 90-110% (slope -3 to -3.5) with R² > 0.98; S-shaped amplification curves with Ct between 15 and 35 (negative control Ct > 35 or no Ct); and a single melt-curve peak. Small Tm differences (within 1°C) between replicates of the same amplicon are normal and acceptable. If diluting the template lowers the Ct, inhibitors or impure template may be present in the undiluted sample. For first-time use, titrate template loading (a 5-10 fold dilution series) and select the amount giving Ct values between 20 and 30; a common starting point is cDNA reverse-transcribed from 500-1000 ng total RNA, diluted 10-fold, using 1 μL per reaction. ROX serves as a passive reference dye that normalizes non-PCR-related fluorescence fluctuations and corrects pipetting and well-to-well variation; incorrectly configured ROX can cause irregular, discontinuous amplification curves. The premix contains stabilizers that do not participate in the reaction and may appear as diffuse smearing on gels; post-amplification gel electrophoresis is not recommended. The reagent is viscous and prone to bubbles: prepare a master mix before aliquoting, dispense slowly along the well wall without introducing air, and briefly centrifuge tubes or gently tap plates to remove residual bubbles. A pale-yellow precipitate after thawing is precipitated stabilizer and does not affect performance; mix until dissolved, or centrifuge and avoid pipetting the precipitate. |
For research use only, not for clinical use.
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