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Biotechnology and Applied Biochemistry (2002) 36, (85–88) (Printed in Great Britain)

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Guide DNA technique in bacterial ribonuclease P reaction for effective processing of tRNA precursor

Terumichi Tanaka¹, Yoshiaki Hori and Yo Kikuchi

Division of Bioscience and Biotechnology, Department of Ecological Engineering, Toyohashi University of Technology, Tempaku-cho, Toyohashi, Aichi 441-8580, Japan

Key words: Escherichia coli, leader sequence, maturation, ribozyme, RNase P.

Abbreviations used: RNase P, ribonuclease P.

¹To whom correspondence should be addressed (e-mail tanakat@eco.tut.ac.jp).

Previously, we found that a small (approx. 20-mer) DNA hybridizing to the 5´-leader region of a tRNA precursor enhances the cleavage efficiency in bacterial ribonuclease P reaction. We named this technique the 'guide DNA technique'. Detailed analyses showed that the length of the guide DNA, concentration of the guide DNA and the hybridizing position affected the cleavage efficiency: for an effective cleavage reaction, guide DNA should be designed to hybridize to the region on the cleavage site, should be 20 bases or more in length and should be of high concentration. The presence of a 5´-flanking region in the DNA did not affect the cleavage reaction. The guide DNA technique is a useful tool for effective preparation of mature tRNA molecules in vitro.

Received 31 May 2002; accepted 11 June 2002

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