CLUE: a bioinformatic and wet-lab pipeline for multiplexed cloning of custom sgRNA libraries.

Becker M, Noll-Puchta H, Amend D, Nolte F, Fuchs C, Jeremias I, Braun CJ (2020)
Nucleic acids research.

Zeitschriftenaufsatz | E-Veröff. vor dem Druck | Englisch
 
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Autor*in
Becker, Martin; Noll-Puchta, Heidi; Amend, Diana; Nolte, Florian; Fuchs, ChristianeUniBi ; Jeremias, Irmela; Braun, Christian J
Abstract / Bemerkung
The systematic perturbation of genomes using CRISPR/Cas9 deciphers gene function at an unprecedented rate, depth and ease. Commercially available sgRNA libraries typically contain tens of thousands of pre-defined constructs, resulting in a complexity challenging to handle. In contrast, custom sgRNA libraries comprise gene sets of self-defined content and size, facilitating experiments under complex conditions such as in vivo systems. To streamline and upscale cloning of custom libraries, we present CLUE, a bioinformatic and wet-lab pipeline for the multiplexed generation of pooled sgRNA libraries. CLUE starts from lists of genes or pasted sequences provided by the user and designs a single synthetic oligonucleotide pool containing various libraries. At the core of the approach, a barcoding strategy for unique primer binding sites allows amplifying different user-defined libraries from one single oligonucleotide pool. We prove the approach to be straightforward, versatile and specific, yielding uniform sgRNA distributions in all resulting libraries, virtually devoid of cross-contaminations. For in silico library multiplexing and design, we established an easy-to-use online platform at www.crispr-clue.de. All in all, CLUE represents a resource-saving approach to produce numerous high quality custom sgRNA libraries in parallel, which will foster their broad use across molecular biosciences. © The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.
Erscheinungsjahr
2020
Zeitschriftentitel
Nucleic acids research
eISSN
1362-4962
Page URI
https://pub.uni-bielefeld.de/record/2943893

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Becker M, Noll-Puchta H, Amend D, et al. CLUE: a bioinformatic and wet-lab pipeline for multiplexed cloning of custom sgRNA libraries. Nucleic acids research. 2020.
Becker, M., Noll-Puchta, H., Amend, D., Nolte, F., Fuchs, C., Jeremias, I., & Braun, C. J. (2020). CLUE: a bioinformatic and wet-lab pipeline for multiplexed cloning of custom sgRNA libraries. Nucleic acids research. doi:10.1093/nar/gkaa459
Becker, M., Noll-Puchta, H., Amend, D., Nolte, F., Fuchs, C., Jeremias, I., and Braun, C. J. (2020). CLUE: a bioinformatic and wet-lab pipeline for multiplexed cloning of custom sgRNA libraries. Nucleic acids research.
Becker, M., et al., 2020. CLUE: a bioinformatic and wet-lab pipeline for multiplexed cloning of custom sgRNA libraries. Nucleic acids research.
M. Becker, et al., “CLUE: a bioinformatic and wet-lab pipeline for multiplexed cloning of custom sgRNA libraries.”, Nucleic acids research, 2020.
Becker, M., Noll-Puchta, H., Amend, D., Nolte, F., Fuchs, C., Jeremias, I., Braun, C.J.: CLUE: a bioinformatic and wet-lab pipeline for multiplexed cloning of custom sgRNA libraries. Nucleic acids research. (2020).
Becker, Martin, Noll-Puchta, Heidi, Amend, Diana, Nolte, Florian, Fuchs, Christiane, Jeremias, Irmela, and Braun, Christian J. “CLUE: a bioinformatic and wet-lab pipeline for multiplexed cloning of custom sgRNA libraries.”. Nucleic acids research (2020).

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