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The Thermo Scientific Maxima First Strand cDNA Synthesis Kit for RT-qPCR is a convenient system optimized for cDNA synthesis in 2-step real time quantitative RT-PCR (RT-qPCR) applications. The kit uses Maxima Reverse Transcriptase (RT), an advanced enzyme derived by in vitro evolution of M-MuLV RT. The enzyme features high thermostability, robustness and increased cDNA synthesis rate compared to wild type M-MuLV RT.
The Maxima First Strand cDNA Synthesis Kit for RT-qPCR is capable of reproducible cDNA synthesis from a wide range of total RNA amounts (1 pg to 5 µg) at elevated temperatures (50 to 65°C).The synthesis reaction can be completed in 15 to 30 minutes. Components of the Maxima First Strand cDNA Synthesis Kit for RT-qPCR are pre-mixed to save time and to reduce the possibility of pipetting errors.
Maxima First Strand cDNA Synthesis Kit for RT-qPCR contains
The recombinant RiboLock RNase Inhibitor, supplied with the kit, effectively protects RNA templates from degradation by RNases A, B and C at temperatures up to 55°C.
Oligo(dT)18 and random hexamer primers are supplied with the kit. Random hexamer primers bind non-specifically and are used to synthesize cDNA from all RNAs in a total RNA population. The oligo(dT)18 primer selectively anneals to the 3’-end of poly(A) RNA, synthesizing cDNA only from poly(A)-tailed mRNA. Gene-specific primers may also be used with the kit to prime synthesis from a specified sequence.
Water, nuclease-free is provided for reaction set-up and dilution of sample RNA. The absence of endo-, exodeoxyribonucleases, ribonucleases and phosphatases has been confirmed by appropriate quality tests.
Only available in US and Canada
The Maxima Universal First Strand cDNA Synthesis Kit differs from the Maxima First Strand cDNA Synthesis Kit by having separate kit components: Maxima Reverse Transcriptase, RiboLock RNase Inhibitor, 5X RT Buffer, 10 mM dNTPs, oligo (dT)18 primer, random hexamer primers, and nuclease-free water.
Highly sensitive two step RT-qPCR.A – amplification plot. B – standard curve.Amplification of the human PPP1CA gene was performed on varying amounts of Jurkat cell total RNA (5 μg, 2.5 μg, 1 μg, 100 ng, 10 ng, 1 ng, 100 pg, 10 pg, 1 pg). First strand cDNA was generated with the Maxima First Strand cDNA Synthesis Kit for RT-qPCR (#K1641). cDNA was amplified with the Maxima Probe /ROX qPCR Master Mix (2X) using the TaqMan assay specific for PPP1CA. Reactions were performed on an ABI® 7500 Real-Time PCR instrument. 1 pg of total RNA was successfully detected. Reaction efficiency was 105%, slope-3.29, R2=0.999.
Reproducible RT-qPCR results using the Maxima First Strand cDNA Synthesis Kit for RT-qPCR.First strand cDNA was generated from 100 ng to 1 pg of total Jurkat cell RNA with the Maxima First Strand cDNA Synthesis Kit for RT-qPCR (#K1641) in 16 replicate reactions. Synthesized cDNA was used as a template in subsequent qPCR with Maxima SYBR Green/ROX qPCR Master Mix (#K0221) on the ABI® 7500 Real-Time PCR instrument. Parallel RT reactions demonstrate reproducible cDNA synthesis and low variability levels with a wide range of starting RNA amounts.
Greater yields and higher sensitivity using Maxima First Strand cDNA Synthesis Kit for RT-qPCR.Amplification of the E. coli 23S RNA gene was performed on 10-fold serial dilutions of total RNA (30 ng to 3 fg). First strand cDNA was generated using the Maxima First Strand cDNA Synthesis Kit for RT-qPCR (red) and a reverse transcription kit from another vendor (green). cDNA was amplified using Maxima SYBR Green/ROX qPCR Master Mix (#K0221) on the ABI® 7500 Real-Time PCR; instrument. All starting RNA amounts (30 ng to 3 fg) were successfully detected using the Maxima First Strand cDNA Synthesis Kit with lower Cq and 102% reaction efficiency.