Comparison of self-sustained sequence-replication reaction systems. NLM AIDSLINE Important note: Information in this article was accurate in 1996. The state of the art may have changed since the publication date.

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Comparison of self-sustained sequence-replication reaction systems.

Eur J Biochem. 1996 Jan 15;235(1-2):256-61. Unique Identifier : AIDSLINE MED/96202944
Gebinoga M; Oehlenschlager F; Institute for Molecular Biotechnology, Jena, Germany.

Abstract: The 3SR (self-sustained sequence-replication) reaction is a very efficient method for isothermal amplification of target DNA or RNA sequences in vitro. This method requires three enzymatic activities: reverse transcriptase, DNA-dependent RNA polymerase and Escherichia coli ribonuclease H. We have modified the original protocol by using human immunodeficiency virus (HIV)-1 reverse transcriptase instead of avian myeloblastosis virus (AMV) reverse transcriptase to allow amplification with T7 RNA polymerase but without E. coli ribonuclease H. Comparison of the incorporation kinetics between the conventional three-enzyme 3SR and our two-enzyme 3SR shows differences in the kinetic behaviour. Furthermore, by the new two-enzyme 3SR, the amplified RNA is obtained in a purer form compared with the experiments with three-enzyme 3SR. The aim of our research is to adapt 3SR as a useful tool for darwinian evolutionary experiments.
Keywords: Base Sequence Comparative Study DNA Primers/GENETICS *DNA Replication Evaluation Studies Fluorescent Dyes Gene Amplification/*METHODS In Vitro Kinetics Molecular Sequence Data Nucleic Acid Conformation Ribonuclease H, Calf Thymus RNA/CHEMISTRY/GENETICS/METABOLISM RNA Polymerases RNA-Directed DNA Polymerase Support, Non-U.S. Gov't Thiazoles Thymine Nucleotides/METABOLISM Uridine Triphosphate/METABOLISM JOURNAL ARTICLEKWDbasesequencecomparativestudydnaprimers/geneticsKWDdnareplicationevaluationstudiesfluorescentdyesgeneamplification/KWDmethodsinvitrokineticsmolecularsequencedatanucleicacidconformationribonucleaseh,calfthymusrna/chemistry/genetics/metabolismrnapolymerasesrna-directeddnapolymerasesupport,non-uKWDsKWDgov'tthiazolesthyminenucleotides/metabolismuridinetriphosphate/metabolismjournalarticle
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M9690916

Copyright © 1996 - National Library of Medicine. Reproduced under license with the National Library of Medicine, Bethesda, MD.

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