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M Gebinoga

Publications and source records attributed to M Gebinoga.

3 recordsLinked to original sources

Cation radius effects on cell-free translation in rabbit reticulocyte lysate.

The effect of monovalent cation concentrations on the translation was examined in the rabbit reticulocyte cell-free system. The translation of standard reporter gene luciferase was studied using different concentrations of LiCl, NaCl, KCl, RbCl, CsCl, NH(4)Cl, and (CH(3))(4)NCl and the acetates of Na(+), K(+), and NH4(+). Only the salts of K(+), Rb(+), and NH4(+) and to some minor extent of Cs(+) significantly supported translation. Optimum concentrations were dependent on the cation used. Optimum concentrations ranged between 40 mM (NH(4)Ac), 80 mM (KCl, NH(4)Cl), and 100 mM (RbCl, KAc). The maximum efficiency of translation depends on the ionic radius of the cation used. KCl and RbCl were superior to all other salts tested in stimulating in vitro translation. The results were confirmed, using a second reporter system, M-hirudin. Here, however, broad optima were observed with RbCl being slightly superior to KCl in supporting translation.

Animals↗

Comparison of self-sustained sequence-replication reaction systems.

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.

Base Sequence↗