ClickSeq: Replacing Fragmentation and Enzymatic Ligation with Click-Chemistry to Prevent Sequence Chimeras

  • Elizabeth Jaworski
  • Andrew RouthEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1712)


We recently reported a fragmentation-free method for the synthesis of Next-Generation Sequencing libraries called “ClickSeq” that uses biorthogonal click-chemistry in place of enzymes for the ligation of sequencing adaptors. We found that this approach dramatically reduces artifactual chimera formation, allowing the study of rare recombination events that include viral replication intermediates and defective-interfering viral RNAs. ClickSeq illustrates how robust, bio-orthogonal chemistry can be harnessed in vitro to capture and dissect complex biological processes. Here, we describe an updated protocol for the synthesis of “ClickSeq” libraries.

Key words

ClickSeq RNAseq Click-chemistry Next-generation sequencing Flock house virus 



This work was supported by UTMB start-up funds and a University of Texas System Rising STARs Award to A.R.


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Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  1. 1.Department of Biochemistry and Molecular BiologyThe University of Texas Medical BranchGalvestonUSA
  2. 2.Sealy Center for Structural Biology and Molecular BiophysicsUniversity of Texas Medical BranchGalvestonUSA

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