Discovery, characterization, and application of chromosomal integration sites in the hyperthermophilic archaeon Sulfolobus islandicus

Boob, Aashutosh Girish; Zhang, Changyi; Pan, Yuwei; Zaidi, Airah; Whitaker, Rachel J.; Zhao, Huimin

doi:10.1016/j.tibtech.2025.11.003

Title: Discovery, characterization, and application of chromosomal integration sites in the hyperthermophilic archaeon Sulfolobus islandicus

Journal Article · Tue Dec 01 00:00:00 UTC 2026 · Trends in Biotechnology

DOI: https://doi.org/10.1016/j.tibtech.2025.11.003 · OSTI ID:3007272

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Univ. of Illinois at Urbana-Champaign, IL (United States)

Sulfolobus islandicus, an emerging archaeal model organism, offers unique advantages for metabolic engineering and synthetic biology applications owing to its ability to thrive in extreme environments. Although several genetic tools have been established for this organism, the lack of well-characterized chromosomal integration sites has limited its potential as a cellular factory. Here, in this work, we systematically identified and characterized 13 artificial CRISPR RNAs targeting eight integration sites in S. islandicus using the CRISPR-COPIES pipeline and a multi-omics-informed computational workflow. We leveraged the endogenous CRISPR-Cas system to integrate the reporter gene lacS and validated heterologous expression through a β-galactosidase assay, revealing significant positional effects. As a proof of concept, we utilized these sites to genetically manipulate lipid ether composition by overexpressing glycerol dibiphytanyl glycerol tetraether (GDGT) ring synthase B (GrsB). This study expands the genetic toolbox for S. islandicus and advances its potential as a robust platform for archaeal synthetic biology and industrial biotechnology.

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Research Organization:: Univ. of Illinois at Urbana-Champaign, IL (United States)

Sponsoring Organization:: National Research Foundation of Korea (NRF); National Science Foundation (NSF); USDOE Office of Science (SC), Biological and Environmental Research (BER)

Grant/Contract Number:: SC0018420

OSTI ID:: 3007272

Journal Information:: Trends in Biotechnology, Journal Name: Trends in Biotechnology Journal Issue: 4 Vol. 44; ISSN 0167-7799

Publisher:: ElsevierCopyright Statement

Country of Publication:: United States

Language:: English

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