The folding of large RNAs studied by hybridization to arrays of complementary oligonucleotides.
Folding pathways of large RNAs are poorly understood. We have addressed this question by hybridizing in vitro transcripts, which varied in size, to an array of antisense oligonucleotides. All transcripts included a common sequence and all but one shared the same start-point; the other had a small deletion of the 5' end. Minimal free energy calculations predicted quite different folds for these transcripts. However, hybridization to the array showed predominant features that were shared by transcripts of all lengths, though some oligonucleotides that hybridized strongly to the short transcripts gave weak interaction with longer transcripts. A full-length RNA fragment that had been denatured by heating and allowed to cool slowly gave the same hybridization result as a shorter transcript. Taken together, these results support theories that RNA folding creates local stable states that are trapped early in the transcription or folding process. As the transcript elongates, interactions are added between regions that are transcribed early and those transcribed late. The method here described helps in identifying regions in the transcripts that take part in long-range interactions.
Sohail
M
M
Department of Biochemistry, University of Oxford, England, United Kingdom. msohail@bioch.ox.ac.uk
Akhtar
S
S
Southern
E M
EM
D006358:Hot Temperature; D046911:Macromolecular Substances; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis; D016376:Oligonucleotides, Antisense / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D012313:RNA / Q000737:chemistry* / Q000378:metabolism
D046911:Macromolecular Substances; D016376:Oligonucleotides, Antisense; D012313:RNA