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Transcriptor First Strand cDNA Synthesis Kit
For robust two-step RT-PCR of RNA up to 14 kb.
Catalog # Pack size
04897030001 1 kit for up to 200 reactions
04896866001 1 kit for up to 100 reactions
04379012001 1 kit for up to 50 reactions, including 10 control reactions
Application
The Transcriptor First Strand cDNA Synthesis Kit is designed to reverse transcribe RNA (mRNA, total RNA, viral RNA, and in vitro-transcribed RNA) from a variety of sources for the following applications:
Studying gene expression levels, via two-step RT-PCR, using qualitative RT-PCR or quantitative RT-PCR on the LightCycler® Carousel-Based System, the LightCycler® 480 System, or other real-time instruments. Generating cDNA libraries with large and full-length inserts.Cloning genes of interest.The kit contains all components required for cDNA reactions for use with conventional thermal cyclers and real-time PCR instruments. In addition, the 50-reaction pack size includes 10 control reactions.
Benefits
Optimized for Many Applications
Use with conventional thermal cyclers and real-time PCR instruments. Optimize real-time PCR – the kit is tested with the LightCycler® Carousel-Based System, the LightCycler® 480 System, and other real-time instruments. Obtain accurate linear quantification over at least a 108 -fold range of input RNA (in vitro transcripts), permitting the analysis of genes with very low or extremely high expression levels. Generate efficient qPCR curves, with high fluorescence intensity and identical distances between RNA dilutions, allowing a straightforward analysis of results (Figure 1). Obtain reliable and sensitive results – there are no additives in the RT buffer system that interfere with – or inhibit – the subsequent PCR.Robustness
Transcribe a variety of templates, even the most difficult (e.g., GC-rich RNA with high secondary structures). Achieve high reproducibility (Figure 2). Generate full-length transcripts with the anchored-oligo(dT)18 primer that is included in the kit. Obtain cDNA transcripts up to 14 kb (Figure 3).Flexibility
Use three different priming methods – random hexamers, anchored-oligo(dT)18, and sequence-specific primers – depending on the type of analysis needed.
Product Description
Transcriptor Reverse Transcriptase – the core component of the kit – is a recombinant reverse transcriptase expressed in E. coli. The enzyme has RNA-directed DNA polymerase activity, DNA-dependent DNA polymerase activity, unwinding activity, and RNase H activity that degrades RNA in RNA:DNA hybrids. The latter circumvents the need to perform an additional time-consuming RNase H incubation step after reverse transcription. This shortens the reaction time and reduces costs.
Transcriptor Reverse Transcriptase is recommended for RT-PCR because of its high sensitivity in combination with high thermostability: the enzyme synthesizes long cDNA products (up to 14 kb) and can be used at temperatures up to 65°C. Due to its thermostability, Transcriptor Reverse Transcriptase is recommended for GC-rich templates with high secondary structure, without the need to include additives in the reaction.
The kit provides all reagents required for first-strand cDNA synthesis reactions. For priming, three different primer systems can be used. Two cDNA synthesis primers are provided with the kit: random hexamer primers and an anchored-oligo(dT)18 primer. The latter is designed to bind at the beginning of the poly(A) tail to generate full-length cDNAs and to prevent priming from internal sites of the poly(A) tail. The 5' ends of long mRNAs are often underrepresented; therefore, this priming method is preferred for most applications. The use of random hexamer primers enables priming thr |