The RT-PCR kit is designed especially for simple and effective one-tube RT-PCR. The reverse transcription step is directed by M-MuLV Reverse Transcriptase, as soon as cDNA is synthesized, a chemically modified “Hot-Start” Taq DNA polymerase, is activated by heating up the reaction 10 min at 95 °C. As soon as the Taq-star enzyme is activated, regular PCR procedure is performed. An optimized reaction buffer provides good performance both for Reverse Transcriptase and for Taq DNA Polymerase. The buffer qA provides good performance in TaqMan assay and in SYBR Green Real Time analysis as well.

General PCR-Conditions

When the reaction mixture is prepared according to the protocol, the PCR cycler can be programmed for synthesis of cDNA as the first step, heating-up step for inactivation of Reverse Transcriptase, simultaneous activation of Taq DNA Polymerase and PCR-cycling for the following steps.

Advantages:

• no additional manipulation after reverse transcription step
• no change of buffers
• no transfer of valuable material
• higher yield of cDNA

Composition

The kit for one tube RT-PCR contains:

• Reaction buffer qA (5x) MgCl ₂ 15 mM
• dNTPs 5 mM
• Mixture of the enzymes
• DNAse-RNAse free water

Storage conditions

-20 °C