BRIEF RESEARCH REPORT article
Front. Bioeng. Biotechnol.
Sec. Synthetic Biology
AistSeq: An In-House Easy-to-purify Tn5-Based Plasmid Sequencing Platform Using A Compact Benchtop Sequencer
Provisionally accepted
- National Institute of Advanced Industrial Science and Technology (AIST), Tokyo, Japan
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Sequence verification of plasmids is a fundamental process in synthetic biology. For plasmid sequence verification using next-generation sequencing (NGS) library preparation, Tn5 transposase is widely used. Streamlined sequencing workflow for laboratory-scale applications is important; however, recombinant Tn5 production in-house can be laborious. In this study, we demonstrated that the addition of a large soluble tag was not essential for purification and that the fusion of a His10 tag and protein A was sufficient to yield active Tn5 transposase in adequate amounts. In addition, we evaluated exonuclease-based genomic DNA digestion for plasmid sequencing from an E. coli lysate and the data analysis pipeline of sequences derived from the Illumina iSeq100 platform for de novo assembly, reference mapping, and annotation. This study proposes a simple workflow of an in-house easy-to-purify Tn5-based plasmid sequencing platform using a compact benchtop sequencer (AistSeq).
Keywords: Tn5 transposase purification, next-generation sequencing (NGS), plasmid sequencing, pipeline, iSeq100
Received: 26 Jul 2025; Accepted: 09 Dec 2025.
Copyright: © 2025 Suzuki, Romsuk, Nakamura, Moriwaki and Sugano. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Shigeo S. Sugano
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