Buvoli A, Buvoli M, Leinwand L A
Department of Molecular, Cellular, and Developmental Biology, University of Colorado at Boulder, 80309, USA.
RNA. 2000 Jun;6(6):912-8. doi: 10.1017/s1355838200000339.
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.
一种通过寡脱氧核糖核苷酸探针杂交极大增强tRNA检测的方法已被开发出来。由于高度结构化的tRNA区域常常妨碍异源双链体的形成，我们测试了一种名为解折叠剂的冷寡脱氧核糖核苷酸破坏tRNA二级/三级结构并促进与反密码子环互补的第二种标记寡核苷酸杂交的能力。在此我们表明，在预杂交/杂交反应中过量的解折叠剂可将几乎检测不到的杂交信号增强200多倍，而不影响探针特异性。这种灵敏的检测方法使得轻松研究和监测tRNA同工受体表达的变化成为可能。
