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Evolutionary dynamics and population control during in vitro selection and amplification with multiple targets

Published online by Cambridge University Press:  07 November 2002

HUA SHI
Affiliation:
Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA
XIAOCHUN FAN
Affiliation:
Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA
ZHUOYU NI
Affiliation:
Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA
JOHN T. LIS
Affiliation:
Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA
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Abstract

Iterative cycles of in vitro selection and amplification allow rare functional nucleic acid molecules, aptamers, to be isolated from large sequence pools. Here we present an analysis of the progression of a selection experiment that simultaneously yielded two families of RNA aptamers against two disparate targets: the intended target protein (B52/SRp55) and the partitioning matrix. We tracked the sequence abundance and binding activity to reveal the enrichment of the aptamers through successive generations of selected pools. The two aptamer families showed distinct trajectories of evolution, as did members within a single family. We also developed a method to control the relative abundance of an aptamer family in selected pools. This method, involving specific ribonuclease digestion, can be used to reduce the background selection for aptamers that bind the matrix. Additionally, it can be used to isolate a full spectrum of aptamers in a sequential and exhaustive manner for all the different targets in a mixture.

Type
Research Article
Copyright
2002 RNA Society

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