Evolution in a Test Tube: Exploring the Structure and Function of RNA Probes
Abstract
This Trident project used the new in vitro selection strategy to explore the structure and function of one of the tobramycin beacon aptamers. The question addressed was the following: What are the sequence/structure constraints on a functional tobramycin beacon aptamer? The sequence of the RNA was partially randomized in order to produce a large number of variants related to the original aptamer. Then, the selection for functional molecules was repeated. The function and the sequences of the RNA molecules were monitored during the course of the selection process. The function of the RNA pools improved after each round of selection until, after nine rounds, the newly selected molecules functioned nearly as well as the original RNA molecules did. An additional five rounds of selection did not result in further improvement. RNAs present at various stages of selection were randomly chosen and sequenced. Many sequence variants remained at the end of the selection, suggesting that there are multiple solutions to the problem of constructing a tobramycin beacon aptamer. The sequence variants were analyzed for clues about structural requirements. Analysis of the sequences variants provided evidence that supports a hypothetical secondary structure. Evidence for additional interactions begins to outline the tertiary structure of the molecule. In addition to increasing our understanding of RNA structure/function relationships, the results of these experiments will aide in the design of future in vitro selection strategies.
Document Details
- Document Type
- Technical Report
- Publication Date
- May 02, 2008
- Accession Number
- ADA486679
Entities
People
- Jeffrey E. Vandenengel
Organizations
- United States Naval Academy