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| Product Name | 1st Strand cDNA Synthesis Kit III (with gDNA Digester) |
| Catalog No. | RESK-0022 |
| Description | The 1st Strand cDNA Synthesis Kit III (with gDNA Digester) is a first-strand cDNA synthesis kit that incorporates a genomic DNA removal step. Built on a third-generation reverse transcriptase, the enzyme synthesizes cDNA rapidly and shows greatly improved thermostability, tolerating reaction temperatures up to 60°C, which makes it ideal for RNA templates with complex secondary structures. Enhanced template affinity makes the kit well suited for small amounts of template and low-copy genes, and its full-length cDNA synthesis capability extends to fragments as long as 19.8 kb. The included gDNA Digester Mix removes residual genomic DNA contamination from RNA templates for more reliable downstream results. Both Random Primers N6 and Oligo (dT)18 are provided, and gene-specific primers can also be used; the resulting single-stranded cDNA is ready for PCR or qPCR. |
| Intended Use | First-strand cDNA synthesis from RNA templates with genomic DNA removal, for downstream PCR or qPCR; suitable for templates with complex secondary structures, low-abundance templates, and low-copy genes. |
| Principle / Technology | Reverse transcription using a thermostable third-generation reverse transcriptase (tolerates up to 60°C) with enhanced template affinity, preceded by genomic DNA digestion with the gDNA Digester Mix. Flexible priming with Random Primers N6, Oligo (dT)18, or gene-specific primers (GSP). |
| Detection Method | Reverse transcription followed by PCR or qPCR. |
| Sample Type | RNA templates from any species, including templates with complex secondary structures, low-input templates, and low-copy genes; compatible with different GC contents and expression abundances. RNA quality, rather than species origin, determines reverse transcription performance. |
| Performance Range / Specifications | Tolerates reaction temperatures up to 60°C; synthesizes cDNA up to 19.8 kb in length; compatible with genes of different GC contents and expression abundances. |
| Sensitivity / LOD | Enhanced template affinity supports reverse transcription of small amounts of template and low-copy genes. |
| Specificity | Flexible primer selection allows high-specificity amplification: Oligo (dT) for full-length cDNA from eukaryotic mRNA, Random Primers for complex structures and prokaryotic templates, and gene-specific primers for known target sequences. |
| Reaction Conditions / Protocol | The 25°C step in the program serves primer annealing (primer binding). Reaction temperatures up to 60°C are tolerated for templates with complex secondary structures. For miRNA stem-loop reverse transcription, replace the kit primers with a designed stem-loop primer (10 μM): 1 μL stem-loop primer, 15 μL reaction mixture from the gDNA removal step, 2 μL 10x Super Buffer, 1 μL RT Enzyme Mix, and 1 μL ddH2O. |
| Components / Formulation | RNase-free H2O: 1 mL (10 T) / 2 x 1 mL (100 T) 5x gDNA Digester Mix: 30 μL (10 T) / 300 μL (100 T) 10x Super Buffer: 20 μL (10 T) / 200 μL (100 T) RT Enzyme Mix: 10 μL (10 T) / 100 μL (100 T) Random Primers N6 (50 μM): 10 μL (10 T) / 100 μL (100 T) Oligo (dT)18 (50 μM): 10 μL (10 T) / 100 μL (100 T) |
| Storage Conditions | Store at -25°C to -15°C, protected from light. |
| Shelf Life | 18 months when stored at -25°C to -15°C, protected from light. |
| Package Specifications | 10 T, 100 T |
| Product Form | Kit |
| Concentration | Random Primers N6: 50 μM; Oligo (dT)18: 50 μM |
| Expiration Date / Stability | Valid for 18 months at -25°C to -15°C, protected from light. |
| Compatibility | Compatible with downstream PCR and qPCR; suitable for genes of different GC contents and expression abundances. With appropriate primer selection, the kit can be applied to lncRNA with poly(A) tails, circRNA, miRNA (with stem-loop primers), and lncRNA without poly(A) tails. |
| Application Notes / Precautions | Primer selection guide: use Oligo (dT) for full-length first-strand cDNA from eukaryotic templates; use Random Primers for targets with complex secondary structures or prokaryotic templates; use gene-specific primers (GSP) when the target sequence is known; for qPCR, Oligo (dT) and Random Primers can be mixed. cDNA for cloning (long products, Oligo dT priming) can be used for qPCR, but quantitative RT products (short fragments from mixed primers) are not recommended for long-fragment conventional PCR. RNA species origin does not limit reverse transcription; RNA quality is the determining factor. Residual RNase in gDNA removal reagents can degrade RNA and impair results; verify RNA integrity by electrophoresis before and after gDNA removal and use RNase-free-grade gDNA removal reagents. For miRNA stem-loop reverse transcription, replace the kit primers with a designed specific stem-loop primer. The 25°C step in the program is for primer binding. |
For research use only, not for clinical use.
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