DETECTION OF RIBONUCLEASE H-GENERATED MESSENGER-RNA FRAGMENTS IN HUMAN LEUKEMIA-CELLS FOLLOWING REVERSIBLE MEMBRANE PERMEABILIZATION IN THE PRESENCE OF ANTISENSE OLIGODEOXYNUCLEOTIDES

被引:94
作者
GILES, RV
SPILLER, DG
TIDD, DM
机构
[1] Department of Biochemistry, University of Liverpool, Liverpool L69 3BX, UK
来源
ANTISENSE RESEARCH AND DEVELOPMENT | 1995年 / 5卷 / 01期
关键词
D O I
10.1089/ard.1995.5.23
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
The involvement of ribonuclease H (RNase H) in antisense phenomena in intact cells has, to date, only been adequately demonstrated for microinjected Xenopus systems, The significance of RNase H for the antisense effects of oligodeoxynucleotides observed in human and other mammalian cell cultures has remained obscure, in part because of inadequate analytic methods. In this report we show that the ''reverse ligation-mediated PCR'' (RL-PCR) procedure permits amplification of RNA fragments produced by oligodeoxynucleotide-directed RNase H activity, We have used this procedure to demonstrate RNase H-dependent antisense effects in irreversibly permeabilized (dead) cells and reversibly permeabilized (live) cells.
引用
收藏
页码:23 / 31
页数:9
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