2'-O-ALKYL OLIGORIBONUCLEOTIDES AS ANTISENSE PROBES

被引:132
作者
IRIBARREN, AM [1 ]
SPROAT, BS [1 ]
NEUNER, P [1 ]
SULSTON, I [1 ]
RYDER, U [1 ]
LAMOND, AI [1 ]
机构
[1] EUROPEAN MOLEC BIOL LAB,W-6900 HEIDELBERG 1,GERMANY
关键词
affinity selection; biotinylation; modified RNA; ribonucleoprotein complexes oligonucleotide synthesis;
D O I
10.1073/pnas.87.19.7747
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
2'-O-Methyl oligoribonucleotides have recently been introduced as antisense probes for studying RNA processing and for affinity purification of RNA-protein complexes. To identify RNA analogues with improved properties for antisense analysis, 2'-O-alkyl oligoribonucleotides were synthesized in which the alkyl moiety was either the three-carbon linear allyl group or the five-carbon branched 3,3-dimethylallyl group. Both these analogues were found to be completely resistant to degradation by either DNA- or RNA-specific nucleases. Use of biotinylated derivatives of the probes to affinity-select ribonucleoprotein particles from crude HeLa cell nuclear extracts showed that the presence of the bulky 3,3-dimethylallyl group significantly reduces affinity selection, whereas the allyl derivative binds rapidly and stably to targeted sequences and affinity-selects efficiently. The allyl derivatives also showed an increase in the level of specific binding to targeted sequences compared with 2'-O-methyl probes of identical sequence. These properties indicate that the 2'-O-allyl oligoribonucleotides are particularly well suited for use as antisense probes.
引用
收藏
页码:7747 / 7751
页数:5
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