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| Product Name | Fast Cell Direct SYBR Green RT-qPCR Kit |
| Catalog No. | RESK-0063 |
| Description | This kit enables direct gene expression analysis from a wide range of cultured animal cells-including adherent and suspension cell lines, primary cultured cells, various stem cells, and iPS cells-without RNA extraction. The streamlined workflow from template preparation through reverse transcription to qPCR analysis can be completed in as little as 1.5 hours, with short hands-on time, simple operation, and a low error rate. The kit includes cell lysis, washing, gDNA removal, reverse transcription, and SYBR Green qPCR detection reagents for effortless gene expression analysis. |
| Intended Use | For direct two-step SYBR Green RT-qPCR gene expression analysis from cultured animal cells without RNA purification (cell lysis + reverse transcription + dye-based qPCR). |
| Principle / Technology | Cultured cells are lysed directly with the FCD Lysis Buffer, washed, and the reaction stopped; genomic DNA is removed with DNase I. The lysate is reverse transcribed with the 4x RT mix, and cDNA is quantified by SYBR Green qPCR using an antibody-based hot-start master mix. |
| Detection Method | SYBR Green dye-based real-time fluorescence detection. |
| Sample Type | Various cultured animal cells, including adherent and suspension cell lines, primary cultured cells, various stem cells, and iPS cells. |
| Performance Range / Specifications | Complete workflow (cell lysis, reverse transcription, and qPCR) in as little as 1.5 hours Amplicon length: 80-300 bp Annealing temperature: 60°C Template GC content: 30%-70% Reverse transcription system tolerates up to 45% cell lysate qPCR system tolerates up to 30% cDNA input Kit stable for 7 days at 4°C; withstands up to 50 freeze-thaw cycles Universal ROX reference dye premixed-no ROX concentration adjustment required |
| Sensitivity / LOD | Sensitive amplification across genes of different GC content (30%, 50%, 70% GC); in tests with adherent and suspension 293 cells, Ct values and reproducibility outperformed competitor products. |
| Specificity | Antibody-based hot start ensures amplification specificity; the DNase I component efficiently digests genomic DNA, outperforming competitor DNase I when challenged with 100-500 ng gDNA input. |
| Reaction Conditions / Protocol | Two-step workflow: 1. Cell lysis with FCD Lysis Buffer, washing with FCD Washing Buffer, reaction stop with FCD Stop Solution, and gDNA removal with DNase I 2. Reverse transcription with 4x FCD RT Mix 3. SYBR Green qPCR with 2x FCD qPCR SYBR Master Mix (recommended annealing temperature: 60°C) |
| Components / Formulation | FCD Lysis Buffer (2 mL / 5 mL) FCD Washing Buffer (8 mL / 20 mL) FCD Stop Solution (100 μL / 250 μL) DNase I (80 μL / 200 μL) 4x FCD RT Mix (200 μL / 500 μL) 2x FCD qPCR SYBR Green Master Mix (2 mL / 5 mL) RNase-Free Water (2 mL / 5 mL) (Volumes shown for the 40 rxns / 100 rxns packages, respectively) |
| Storage Conditions | Store unopened FCD Lysis Buffer and FCD Washing Buffer at -25 to -15°C; once thawed, keep at 4°C and avoid contamination. Store all other components long-term at -25 to -15°C. |
| Shelf Life | 6 months. |
| Package Specifications | 40 rxns; 100 rxns |
| Product Form | Mix |
| Concentration | 4x (RT mix); 2x (qPCR master mix) |
| Expiration Date / Stability | The kit is stable for 7 days at 4°C and tolerates up to 50 freeze-thaw cycles. cDNA products remain stable (ΔCT within ±0.5) after storage at -20°C, 4°C, 25°C, or 37°C for up to 14 days. |
| Compatibility | Universal ROX reference dye premixed; compatible with the vast majority of real-time fluorescence quantitative PCR instruments without ROX adjustment. |
| Application Notes / Precautions | This kit uses SYBR Green dye-based detection; a probe-based (TaqMan) version of the kit is available for probe-based detection workflows. |
For research use only, not for clinical use.
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