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Reverse transcriptases suitable for one-step RT-PCRb

Product size Type of primers Primer anneal. temp. Sensitivity Specifi-
city
Fidelity in PCR Tolerance
of non-
specific RNA
Tolerance
of serum components
Carry-over prevention dUTP/UNGc Product to select mRNA Viral RNA/RNA rich in secondary structure up to 1kb specific 55-70C + + + - + + + Tth DNA Polymerase up to 3kb specific 55-70C ++ ++ ++ + +++ ++ + C. therm. Polymerase One-step RT-PCR Systemh up to 6kb specific 45-60C +++ + + ++ ++ + not suitable Titan One Tube RT-PCR System/Kit a For more information on preparing total RNA, mRNA, and viral RNA, please visit our Nucleic Acid Isolation and Purification Special In terest 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 comparatively 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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