De novo assembly and next-generation sequencing to analyse full-length gene variants from codon-barcoded libraries

Namjin Cho, Byungjin Hwang, Jung-ki Yoon, Sangun Park, Joongoo Lee, Han Na Seo, Jeewon Lee, Sunghoon Huh, Jinsoo Chung and Duhee Bang ()
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Namjin Cho: Yonsei University
Byungjin Hwang: Yonsei University
Jung-ki Yoon: College of Medicine, Seoul National University
Sangun Park: Korea Military Academy
Joongoo Lee: University of Oxford
Han Na Seo: Yonsei University
Jeewon Lee: Yonsei University
Sunghoon Huh: Yonsei University
Jinsoo Chung: University of California
Duhee Bang: Yonsei University

Nature Communications, 2015, vol. 6, issue 1, 1-9

Abstract: Abstract Interpreting epistatic interactions is crucial for understanding evolutionary dynamics of complex genetic systems and unveiling structure and function of genetic pathways. Although high resolution mapping of en masse variant libraries renders molecular biologists to address genotype-phenotype relationships, long-read sequencing technology remains indispensable to assess functional relationship between mutations that lie far apart. Here, we introduce JigsawSeq for multiplexed sequence identification of pooled gene variant libraries by combining a codon-based molecular barcoding strategy and de novo assembly of short-read data. We first validate JigsawSeq on small sub-pools and observed high precision and recall at various experimental settings. With extensive simulations, we then apply JigsawSeq to large-scale gene variant libraries to show that our method can be reliably scaled using next-generation sequencing. JigsawSeq may serve as a rapid screening tool for functional genomics and offer the opportunity to explore evolutionary trajectories of protein variants.

Date: 2015
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DOI: 10.1038/ncomms9351

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