Amplification and Re-Generation of LNA-Modified Libraries

H. Doessing; L. Hansen; R. Veedu; J. Wengel; B. Vester

doi:10.3390/molecules171113087

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Amplification and Re-Generation of LNA-Modified Libraries

Journal article

Open access

Peer reviewed

Amplification and Re-Generation of LNA-Modified Libraries

H. Doessing, L. Hansen, R. Veedu, J. Wengel and B. Vester

Molecules, Vol.17(12), pp.13087-13097

2012

DOI: https://doi.org/10.3390/molecules171113087

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Abstract

Locked nucleic acids (LNA) confer high thermal stability and nuclease resistance to oligonucleotides. The discovery of polymerases that accept LNA triphosphates has led us to propose a scheme for the amplification and re-generation of LNA-containing oligonucleotide libraries. Such libraries could be used for in vitro selection of e.g., native LNA aptamers. We maintained an oligonucleotide library encoding 40 randomized positions with LNA ATP, GTP, CTP, and TTP for 7 rounds of ‘mock’ in vitro selection in the absence of a target and analyzed the sequence composition after rounds 1, 4 and 7. We observed a decrease in LNA-A content from 20.5% in round 1 to 6.6% in round 7. This decrease was accompanied by a substantial bias against successive LNA-As (poly-LNA adenosine tracts) and a relative over-representation of single LNA-As. Maintaining a library with LNA TTP yielded similar results. Together, these results suggest that dispersed LNA monomers are tolerated in our in vitro selection protocol, and that LNA-modified libraries can be sustained for up to at least seven selection rounds, albeit at reduced levels. This enables the discovery of native LNA aptamers and similar oligonucleotide structures.

Details

Title: Amplification and Re-Generation of LNA-Modified Libraries
Authors/Creators: H. Doessing (Author/Creator) - University of Southern Denmark
L. Hansen (Author/Creator) - Institut for Biokemi og Molekylær Biologi
R. Veedu (Author/Creator) - NAC, Institut for Fysik og Kemi
J. Wengel (Author/Creator) - University of Southern Denmark
B. Vester (Author/Creator) - Institut for Biokemi og Molekylær Biologi
Publication Details: Molecules, Vol.17(12), pp.13087-13097
Publisher: MDPI
Identifiers: 991005543188907891
Copyright: © 2012 MDPI AG
Murdoch Affiliation: Murdoch University
Language: English
Resource Type: Journal article

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Collaboration types: Domestic collaboration; International collaboration
Citation topics: 2 Chemistry; 2.145 Biosensors; 2.145.243 Nanobiosensors
Web Of Science research areas: Biochemistry & Molecular Biology; Chemistry, Multidisciplinary
ESI research areas: Chemistry