Transcriptor High Fidelity cDNA Synthesis Kit

Descrição detalhada : Retroviral reverse transcriptases commonly used for cDNA synthesis exhibit a higher error rate than other DNA polymerases used in nucleic acid analysis techniques. This lack in accuracy leads to a significant number of base exchanges or frameshifts, which are further propagated in subsequent PCR reactions. High fidelity (proofreading) PCR enzymes have been available for many years; Roche's new high accuracy reverse transcriptase is further able to synthesize high yields of full-length cDNA.High mutation rate is a hallmark of retrovirus replication. This originates in the mechanism of genome replication by the viral-encoded reverse transcriptase, which converts the genomic RNA of the virus to a dsDNA. During this process, reverse transcription produces frequent replication errors. One accepted explanation of this inaccuracy is the lack of RT 3'-5' exonuclease activity. The naturally high error rate of reverse transcriptases is not optimal for many different applications., Transcriptor High Fidelity Reverse Transcriptase enzyme blend for high fidelity two-step RT-PCR of RNA up to 14 kb.The core component of the Transcriptor High Fidelity Synthesis cDNA Synthesis kit is the Transcriptor High Fidelity Reverse Transcriptase, a blend of a recombinant reverse transcriptase and a proofreading mediating enzyme. The synergy between both enzymes is the key to the ability of the enzyme blend to reverse transcribe RNA templates with 7-fold higher fidelity compared to other commonly used reverse transcriptases.The Transcriptor High Fidelity Reverse Transcriptase enzyme blend efficiently reverse transcribes templates up to 14 kb. Due to the high thermostability of both enzyme components and the specially optimized buffer system, reverse transcription is possible at temperatures up to +55C. This allows the reverse transcription of GC-rich templates with high secondary structure, without the need to include additives that may negatively influence the accuracy of the reverse transcription reaction.The recombinant reverse transcriptase included in the enzyme blend is 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, reducing reaction time and costs.The Transcriptor High Fidelity cDNA Synthesis 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 throughout the length of RNA for uniform representation of all RNA sequences and allows reverse transcription of RNAs that do not carry a poly(A) tail. The thermostable Protector RNase Inhibitor (included in the kit) protects RNA from degradation at high reaction temperatures.
Sinónimos : RT-PCR; cDNA sysnthesis
Armazenamento : -20°C to 0°C
Embalagem : 1X1EA