Aptamers targeting RNA molecules.
Oligonucleotides complementary to RNA sequences interact poorly with folded target regions. In vitro selection of oligonucleotides carried out against RNA structures have led to aptamers that frequently differ from antisense sequences, but rather take advantage of non-double-stranded peculiarities of the target. Studies along this line provide information about tertiary RNA architectures as well as their interaction with ligand of interest. We describe here a genomic SELEX approach and its application to the recognition of stem-loop structures prone to the formation of kissing complexes. We also provide technical details for running a procedure termed 2D-SELEX that takes advantage of both in vitro selection and dynamic combinatorial chemistry. This allows selecting aptamer derivatives containing modified nucleotides that cannot be incorporated by polymerases. Last we present in vitro transcription conditions under which large amounts of RNA, suitable for NMR structural studies, can be obtained. These different aspects of the SELEX technology have been applied to the trans-activating responsive element of the human immunodeficiency virus type 1, which is crucial for the transcription of the retroviral genome.
Watrin
Marguerite
M
Institut Européen de Chimie et Biologie, Pessac, France, Université Victor Segalen, Bordeaux, France.
Dausse
Eric
E
Lebars
Isabelle
I
Rayner
Bernard
B
Bugaut
Anthony
A
Toulmé
Jean-Jacques
JJ
D052157:Aptamers, Nucleotide / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D020650:Combinatorial Chemistry Techniques; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000235:genetics; D006801:Humans; D055029:Inverted Repeat Sequences; D019906:Nuclear Magnetic Resonance, Biomolecular; D009690:Nucleic Acid Conformation; D012313:RNA / Q000737:chemistry* / Q000378:metabolism*; D052156:SELEX Aptamer Technique / Q000379:methods*
D052157:Aptamers, Nucleotide; D012313:RNA