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| Product Name | 1st Strand cDNA Synthesis SuperMix III for qPCR (with gDNA Digester) |
| Catalog No. | RESK-0018 |
| Description | 1st Strand cDNA Synthesis SuperMix III for qPCR (with gDNA Digester) is a ready-to-use reverse transcription premix built on a third-generation, highly thermostable reverse transcriptase. Compared with the previous generation, the enzyme offers dramatically improved thermal stability—tolerating reaction temperatures up to 60°C—making it ideal for RNA templates with complex secondary structures. Its enhanced template affinity also makes it well suited for low-abundance templates and low-copy-number genes. The kit includes a 5× gDNA Digester Mix that removes residual genomic DNA contamination from RNA templates for more reliable downstream results, and a 4× SuperMix plus containing every component needed for reverse transcription (buffer, dNTPs, reverse transcriptase, RNase inhibitor, and a ratio-optimized Random Primers/Oligo(dT)18 primer mix). Simply add RNA template and RNase-free water; the reverse transcription reaction simultaneously inactivates the gDNA digester, preserving cDNA integrity. Optimized for two-step RT-qPCR, the balanced primer mix initiates cDNA synthesis from all regions of RNA transcripts with equal efficiency, maximizing the authenticity and reproducibility of qPCR results. The cDNA product is compatible with both SYBR Green and probe-based qPCR. |
| Intended Use | First-strand cDNA synthesis with removal of residual genomic DNA for two-step RT-qPCR gene expression analysis; cDNA product is compatible with SYBR Green and probe-based qPCR. Conventional-length gene cloning applications are also possible. |
| Principle / Technology | The premix is based on a thermostable reverse transcriptase that withstands reaction temperatures up to 60°C, allowing reverse transcription of RNA templates with complex secondary structures, and features enhanced template affinity for low-input templates and low-copy genes. A 5× gDNA Digester Mix enzymatically removes residual genomic DNA from the RNA template with no separate inactivation step required—the subsequent reverse transcription reaction terminates digester activity to protect cDNA integrity. A ratio-optimized Random Primers/Oligo(dT)18 primer mix ensures uniform cDNA synthesis efficiency across all regions of RNA transcripts, supporting accurate and reproducible qPCR quantification. |
| Detection Method | Downstream fluorescent quantitative PCR (SYBR Green or probe-based) |
| Sample Type | Total RNA (10 pg–5 μg), including high-GC templates, templates with complex secondary structures, low-abundance templates, and low-copy-number gene targets |
| Performance Range / Specifications | Thermostable reverse transcriptase tolerates reaction temperatures up to 65°C, suitable for high-GC and structurally complex RNA templates. Higher reverse transcription efficiency than comparable market products across genes of different GC content and expression abundance. Broad linear detection range: efficient cDNA synthesis over total RNA inputs of 10 pg–5 μg. RNA input volume of up to 12 μL per reaction, facilitating detection of low-concentration templates. Efficient removal of residual genomic DNA with no additional inactivation step. Both gDNA removal and reverse transcription reagents are supplied as mixes for convenient pipetting with minimal error. |
| Sensitivity / LOD | Efficient cDNA synthesis from as little as 10 pg of total RNA; enhanced template affinity and up to 12 μL RNA input volume support detection of low-concentration templates. |
| Specificity | The included 5× gDNA Digester Mix efficiently removes residual genomic DNA contamination from RNA templates, ensuring downstream qPCR results originate from cDNA and improving reliability. |
| Reaction Conditions / Protocol | Set the reverse transcription temperature at 55°C for routine templates; for templates with complex secondary structures or high GC content, raise the temperature up to 65°C to improve reverse transcription efficiency. The gDNA digestion step may be omitted if genomic DNA has already been removed during RNA extraction. |
| Components / Formulation | RNase-free H2O 5× gDNA Digester Mix 4× SuperMix plus (containing buffer, dNTPs, thermostable reverse transcriptase, RNase inhibitor, and Random Primers/Oligo(dT)18 primer mix) |
| Storage Conditions | Store at -20°C. |
| Shelf Life | 18 months when stored at -20°C. |
| Package Specifications | 10 T 100 T |
| Quality Control | Reverse transcription efficiency benchmarked against comparable market products across genes of different GC content and expression abundance, demonstrating higher efficiency. |
| Shipping Conditions | Shipped with ice packs. |
| Expiration Date / Stability | Valid for 18 months when stored at -20°C. |
| Compatibility | The cDNA product is compatible with SYBR Green and probe-based qPCR; pair with Alta DiagnoTech's qPCR SYBR Green master mixes or qPCR TaqMan probe master mix according to the experimental objective for high-performance gene expression analysis. |
| Application Notes / Precautions | Recommended for qPCR experiments; conventional-length gene cloning is also supported. The kit contains only 3 components, each pipetted at volumes above 1 μL, minimizing pipetting error and improving reproducibility. Use 55°C for routine templates; increase to up to 65°C for templates with complex secondary structures or high GC content. The gDNA removal step can be omitted when genomic DNA has already been eliminated during RNA extraction. For low-concentration templates, increase the RNA input volume (up to 12 μL) to raise the template amount and improve detection. |
For research use only, not for clinical use.
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