QIAGEN OneStep RT-PCR

The QIAGEN OneStep RT-PCR Kit is designed for easy and sensitive one-step RT-PCR using any RNA template. A unique enzyme combination and specially developed reaction buffer ensure efficient reverse transcription and PCR in one tube.

Features and benefits

• Easy one-tube setup
• One-step RT-PCR of any RNA template without optimization
• Unique blend of Omniscript and Sensiscript RTs and HotStarTaq DNA Polymerase for high specificity and sensitivity
• Optimized buffer for efficient reverse transcription and amplification

Principle

QIAGEN OneStep RT-PCR Enzyme Mix contains a specially formulated enzyme blend for both reverse transcription and PCR. The high affinity and wide dynamic range provided by a unique combination of Omniscript and Sensiscript Reverse Transcriptases ensures highly specific and sensitive reverse transcription (see figure "Efficient and Sensitive RT-PCR"). In the same enzyme mix, HotStarTaq DNA Polymerase provides a hot start for high specificity in PCR without compromising reverse transcription.

QIAGEN OneStep RT-PCR Buffer,* in combination with the high-affinity reverse transcriptases, allows effective reverse transcription at higher temperatures to resolve RNA secondary structures. As with all QIAGEN PCR buffers, the QIAGEN OneStep RT-PCR Buffer also provides the advantage of a balanced combination of cations to ensure specific primer annealing without optimization (see figure "Specificity Over a Wide Temperature Range").

* Patent pending.

Efficient and Sensitive RT-PCR

RT-PCR was carried out with HeLa cell total RNA using different one-step RT-PCR kits according to suppliers’ specification. A 311 bp β-actin fragment was amplified using either  A  10 ng template RNA and 30 PCR cycles or  B  10 pg template RNA and 40 PCR cycles. M: markers.

Specificity Over a Wide Temperature Range

RT-PCR was carried out with HeLa cell total RNA using the QIAGEN OneStep RT-PCR Kit or the one-step RT-PCR kit from Supplier AII. RT-PCR was carried out according to suppliers’ specifications, changing only the PCR annealing temperature in 2.5°C increments, as indicated. A 690 bp fragment of the human α-catenin gene was amplified using 10 ng template RNA and 30 PCR cycles. M: markers.

Procedure

The QIAGEN OneStep RT-PCR Kit allows fast, simple, and sensitive one-step RT-PCR with any RNA template. The fast and easy setup combines gene-specific primers and all reaction components in one tube. Nothing needs to be added once the reaction starts (see flowchart).

Applications

The QIAGEN OneStep RT-PCR Kit is suitable for RT-PCR applications such as:

• Gene-expression analysis
• Virus detection
• Single-cell RT-PCR

QIAGEN OneStep RT-PCR Kit

For easy and sensitive one-step RT-PCR

操作手册:QIAGEN OneStep RT-PCR Kit Handbook 05/02 (PDF version, 172 KB)