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Reverse transcriptases suitable for two-step RT-PCRd

Product size Type of primers Primer anneal. temp. Sensitivitye Specificityf Fidelity in RT GC-rich structures Product to selectg total RNA viral RNA specific primer oligo dt primer up to 1kb specific 55-70C n.d. n.d. n.d. n.d. - n.d. Tth DNA Polymerase up to 4kb specific, oligo(dT) hexamers 55-70C + + (+) ++ ++ ++ +++ C. therm. Polymerase for RT in Two-step RT-PCRh up to 10kb specific, oligo(dT) hexamers 37C + + + n.d. + + M-MuLV Reverse Transcriptase up to 12kb specific, oligo(dT)

hexamers 42C (up to 60C) ++

++ ++

++ +

+ +

+ +

+ +

+ AMV Reverse Transcriptase First Strand cDNA Synthesis Kit for RT-PCR (AMV) up to 14kb specific, oligo(dT) hexamers 42C (up to 50C) ++ ++ + + + + Expand Rverse Transcriptasei a For more information on preparing total RNA, mRNA, and viral RNA, please visit our Nucleic Acid Isolation and Purification Special Interest Site b One-step RT-PCR offers faster reactions, reduced risk of contamination, and improved sensitivity. c For prevention of carry-over contamination with Uracil-DNA Glycosylase (UNG) during One-Step RT-PCR, we recommend using the C. therm. DNA Polymerase One-Step RT-PCR System for products up to 3 kb. For short products (up to 1 kb), you may also use the Tth DNA Polymerase. However note that Tth DNA Polymerase is partially inhibited when using dUTP/UNG instead of dTTP. d Two-step RT-PCR offers maximum flexibility with respect to choice of primers, enzyme, and temperature. Two-step RT-PCR also allows the product of a single reverse transcription to be used in several PCR reactions (for analysis of multiple transcripts). e Sensitivity was com paratively tested for fragments in the range of 0.13.0 kb. f Reverse transcription was performed with a sequence specific primer which was then also used for the subsequent PCR step. g Each of the products in this column should be used for first-strand cDNA synthesis only (first step). h C. therm. Polymerase yields optimal results with GC-rich RNA templates. i Not for sale in the U.S.


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