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| Product Name | Oligo dT Primer (18-mer) for cDNA Synthesis |
| Catalog No. | PRIPRO-0020 |
| Description | A pre-synthesized oligonucleotide consisting of 18 consecutive deoxythymidine residues (dT₁₈), designed for selective priming of polyadenylated (poly-A) mRNA during first-strand cDNA synthesis. Oligo-dT primer hybridizes specifically to the 3′ poly-A tail of eukaryotic mRNAs, ensuring that reverse transcription is initiated near the 3′ end of intact transcripts. This targeted priming selectively reverse transcribes mRNA while excluding ribosomal RNA (rRNA) and transfer RNA (tRNA), enriching the resulting cDNA population for protein-coding transcripts. The 18-mer length provides optimal balance between hybridization specificity and melting temperature (Tm ~35–40 °C under typical RT buffer conditions). The product is desalted or HPLC purified and supplied as a ready-to-use solution. |
| Intended Use | Designed for selective first-strand cDNA synthesis from eukaryotic poly-A mRNA in reverse transcription applications including qPCR gene expression analysis, full-length cDNA library construction, 3′-RACE (rapid amplification of cDNA ends), and cDNA cloning. |
| Principle / Technology | Sequence-specific hybridization of dT₁₈ to the poly-A tail of mature eukaryotic mRNA; reverse transcriptase extends from the hybridized primer, generating cDNA from the 3′ end of transcripts |
| Detection Method | cDNA synthesis by reverse transcriptase; oligo-dT priming selectively enriches for polyadenylated RNA; specificity validated by rRNA depletion in synthesized cDNA |
| Sample Type | Total RNA from eukaryotic cells, tissues, or organisms; performs best with intact RNA (RIN ≥7); not suitable for prokaryotic RNA or degraded RNA lacking intact poly-A tails |
| Performance Range / Specifications | 18-mer poly-dT; Tm ~35–40 °C in standard RT buffer; primes eukaryotic mRNA with 3′ poly-A tail of ≥12 adenosine residues; effective for RNAs with poly-A length ≥20 nt |
| Sensitivity / LOD | Enables cDNA synthesis from as low as 10 pg of purified mRNA or 100 pg total RNA; approximately 1–5% of total RNA is mRNA in most eukaryotic samples |
| Specificity | Highly selective for polyadenylated RNA; negligible priming of rRNA and tRNA; cDNA product enrichment for mRNA typically >80% versus random priming which generates >80% rRNA-derived cDNA |
| Reaction Conditions / Protocol | Use 0.5–1 µg (25–50 pmol) oligo-dT₁₈ per 20 µL RT reaction. Incubate with RNA at 65 °C for 5 min, chill on ice, add RT mix. Prime at 42 °C for 30–60 min. Inactivate at 70 °C for 15 min. |
| Components / Formulation | Lyophilized oligo-dT₁₈ or pre-dissolved at 500 µg/mL (~85 µM) in nuclease-free water |
| Storage Conditions | –20 °C; stable in solution; minimize freeze-thaw cycles |
| Shelf Life | 24 months at –20 °C; 12 months at 2–8 °C |
| Package Specifications | Tube containing 5–50 µg; supplied lyophilized or as ready-to-use solution |
| Product Form | Lyophilized white powder or clear colorless solution |
| Quality Control | HPLC or PAGE purity ≥90%; MALDI-TOF MS confirming 18-mer mass; functional test: cDNA synthesis from 1 µg HeLa total RNA with GAPDH qPCR Ct ≤20; absence of DNase and RNase activity. |
| Key Features | Selective mRNA priming excludes rRNA/tRNA; generates full-length cDNA from intact transcripts; ideal for 3′-biased gene expression analysis; compatible with all RT enzymes; economical and reliable; ready-to-use solution available. |
| Purity | ≥90% by HPLC or PAGE; single-length product (18-mer) |
| Concentration | 500 µg/mL (~85 µM) ready-to-use or lyophilized for custom resuspension |
| Activity / Unit Definition | Hybridizes to poly-A tails at 37–42 °C; efficiently primes MMLV, AMV, and SuperScript reverse transcriptases; full-length cDNA >5 kb achievable with optimized conditions |
| Molecular Weight | ~5,500 Da (18 dT residues; approximately 305 Da per dT) |
| Source / Origin | Synthetic; dT phosphoramidite chemistry; no biological raw materials or templates |
| pH Range / Optimal pH | Stable and functional in RT buffer pH 7.5–8.5 |
| Shipping Conditions | Ambient temperature (lyophilized) or on dry ice (solution); oligo-dT is highly stable |
| Expiration Date / Stability | 24 months at –20 °C; 12 months at 2–8 °C; working aliquots stable for several months at –20 °C |
| Regulatory / Compliance | For research use only; not for diagnostic procedures |
| Compatibility | Compatible with all commercial reverse transcriptases: SuperScript II/III/IV, Maxima H Minus, PrimeScript, MMLV, AMV; compatible with all commercial cDNA synthesis kits offering oligo-dT priming |
| Recommended Buffer System | Nuclease-free water or TE buffer; compatible with all commercial RT buffer formulations |
| Application Notes / Precautions | For 3′ RACE, use oligo-dT with an anchor sequence at the 5′ end (anchor-dT primer). For RNA with limited poly-A, supplement with random hexamers for more complete coverage. Avoid using oligo-dT with formalin-fixed or degraded RNA samples — use random hexamers instead. |
| Batch-to-Batch Consistency | Length verified by MS: 18-mer ±0 residues across batches; cDNA synthesis efficiency within ±15% by GAPDH Ct; concentration ±10% |
For research use only, not for clinical use.
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