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Sensiscript RT Kit

For reverse transcription of less than 50 ng RNA per reaction for end-point PCR

  • Sensitive detection of as few as 10 copies of template
  • High cDNA yields due to high-affinity enzyme
  • No additional RNase H digestion step required
  • Fast and easy procedure with no tedious pipetting steps
The Sensiscript Reverse Transcriptase (RT) Kit is ideally suited for highly sensitive applications using very small amounts of RNA (less than 50 ng), such as single-cell RT-PCR and analysis of biopsies and LMD samples. Sensiscript Reverse Transcriptase allows highly efficient RT-PCR over a wide dynamic range and extremely sensitive RT-PCR with very small RNA amounts due to its high affinity for RNA. The Sensiscript RT Kit includes Sensiscript Reverse Transcriptase, dNTPs, and an optimized reaction buffer. Simply add primers for fast and easy cDNA synthesis.

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Cat No./ID: 205211
Sensiscript RT Kit (50)
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For 50 reverse-transcription reactions: Sensiscript Reverse Transcriptase, 150 µl 10x Buffer RT, 100 µl dNTP Mix (contains 5 mM each dNTP), 1.1 ml RNase-free water
Cat No./ID: 205213
Sensiscript RT Kit (200)
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For 200 reverse-transcription reactions: Sensiscript Reverse Transcriptase, 4 x 150 µl 10x Buffer RT, 4 x 100 µl dNTP Mix (contains 5 mM each dNTP), 4 x 1.1 ml RNase-free water
The Sensiscript RT Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

Product Details

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Sensiscript Reverse Transcriptase procedure.

The Sensiscript RT Kit includes dNTPs and an optimized reaction buffer that, together with the high affinity of Sensiscript Reverse Transcriptase for RNA, enables read-through of templates with high GC content or complex secondary structures. Due to this pre-optimization, tedious pipetting and pre-incubation steps are eliminated, and no additional RNase H digestion step is required.

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RT-PCR with RNA corresponding to 1 cell.

Total RNA was isolated from 10 to 5000 HeLa cells using RNeasy technology. One-tenth of each eluate (corresponding to RNA from 1 to 500 cells) was used for reverse transcription with Sensiscript Reverse Transcriptase. One-half of each RT reaction was then used in a 40-cycle PCR with QIAGEN Taq DNA Polymerase and primers specific for genes encoding β-actin, NF-κB, inositol-1,4,5-triphosphate receptor, or dystrophin. Sizes of the amplicons are as indicated.

Performance

Sensiscript Reverse Transcriptase (RT) provides outstanding efficiency over a wide dynamic range and high sensitivity for very small amounts of RNA (see figure "RT-PCR with RNA corresponding to 1 cell"). Regions of RNA with high GC content can cause other reverse transcriptases to stop, dissociate from the RNA template, or skip over looped-out regions of RNA (see figure Full-length RT-PCR products — B). These difficult templates, however, prove no problem for QIAGEN reverse transcriptases (see figure Full-length RT-PCR products — A). With no need for optimization, the Sensiscript RT Kit makes reverse transcription virtually trouble-free.

Lot-to-lot reproducibility of Sensiscript RT Kits is ensured by rigorous quality control at QIAGEN. The optimized Buffer RT, dNTPs, and water included in all Sensiscript RT Kits are guaranteed RNase-free, and each lot of Sensiscript RT is thoroughly tested for RT-PCR reproducibility.

Principle

Sensiscript Reverse Transcriptase (RT) is the first commercially available enzyme designed for highly sensitive applications with very small amounts of RNA (<50 ng), such as single-cell RT-PCR or analysis of small biopsies. It is provided ready-to-use with dNTPs and in an optimized reaction buffer that, together with the high affinity of Sensiscript RT for RNA, results in highly specific and sensitive RT-PCR, even with low-copy transcripts, and the ability to read through even complex RNA secondary structures without adjusting temperature or reaction conditions.  Please note that the primer mix is not provided.

For viral RNA, Omniscript RT is usually the enzyme of choice due to the presence of carrier RNA in most viral RNA preparations.

Procedure
With the optimized Sensiscript Reverse Transcriptase reaction buffer, tedious pipetting and pre-incubation steps are eliminated, and no additional RNase H digestion step is needed (see flowchart "Sensiscript Reverse Transcriptase Procedure").
Applications

Sensiscript Reverse Transcriptase is suitable for use in the following applications:

  • Standard RT and RT-PCR
  • Quantitative, real-time RT-PCR
  • Differential display RT-PCR
  • Primer extension and RACE analysis
  • Synthesis of double-stranded cDNA for cloning

Specifications

Features
Specifications
Applications cDNA synthesis, RT-PCR, qRT-PCR
Enzyme activity Reverse transcription
Mastermix No
Reaction type Reverse transcription
Real-time or endpoint Endpoint
Sample/target type RNA template
Single or multiplex Single
With/without hotstart Without hotstart

Product Resources

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Kit Handbooks (1)
(EN) - Sensiscript Reverse Transcription Handbook
Sensiscript Reverse Transcriptase for First-strand cDNA synthesis using <50 ng RNA; Two-tube RT-PCR; One-tube RT-PCR
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User-Developed Protocols (1)
Labeling of cDNA using labeled dCTP and <50 ng RNA with the Sensiscript™ RT Kit (Labeling protocol C-50) - (EN)
The protocol has been used successfully for Cy3-, Cy5-, and biotin-labeling of cDNA from <50 ng of total RNA or poly A+ mRNA.
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