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Enhanced detection of tRNA isoacceptors by combinatorial oligonucleotide hybridization

Published online by Cambridge University Press:  01 June 2000

ADA BUVOLI ,
MASSIMO BUVOLI  and
LESLIE A. LEINWAND
ADA BUVOLI
Affiliation:
Department of Molecular, Cellular, and Developmental Biology, University of Colorado at Boulder, Boulder, Colorado 80309, USA
MASSIMO BUVOLI
Affiliation:
Department of Molecular, Cellular, and Developmental Biology, University of Colorado at Boulder, Boulder, Colorado 80309, USA
LESLIE A. LEINWAND
Affiliation:
Department of Molecular, Cellular, and Developmental Biology, University of Colorado at Boulder, Boulder, Colorado 80309, USA
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Abstract

A method that greatly enhances the detection of tRNA by oligodeoxyribonucleotide probe hybridization has been developed. Because highly structured tRNA regions often preclude heteroduplex formation, we have tested the ability of cold oligodeoxyribonucleotides called unfolders to disrupt the tRNA secondary/tertiary structures and promote hybridization of a second labeled oligonucleotide complementary to the anticodon loop. Here we show that an excess of unfolders in the pre/hybridization reaction can enhance a barely detectable hybridization signal by more than 200-fold without affecting probe specificity. This sensitive assay makes it possible to easily study and monitor changes in tRNA isoacceptor expression.

Keywords

isoacceptorsoligodeoxyribonucleotide hybridizationtRNA secondary/tertiary structureunfolders
Type
METHOD
Information
RNA , Volume 6 , Issue 6 , June 2000 , pp. 912 - 918
Copyright
2000 RNA Society

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