دورية أكاديمية
أعرض في EDS

Genetic toolbox for Photorhabdus and Xenorhabdus: pSEVA based heterologous expression systems and CRISPR/Cpf1 based genome editing for rapid natural product profiling.

التفاصيل البيبلوغرافية
العنوان: Genetic toolbox for Photorhabdus and Xenorhabdus: pSEVA based heterologous expression systems and CRISPR/Cpf1 based genome editing for rapid natural product profiling.
المؤلفون: Rill A; Department of Natural Products in Organismic Interactions, Max-Planck Institute for Terrestrial Microbiology, 35043, Marburg, Germany.; Molecular Biotechnology, Department of Biosciences, Goethe University Frankfurt, 60438, Frankfurt am Main, Germany.; Department of Chemistry, Chemical Biology, Phillips University Marburg, 35043, Marburg, Germany., Zhao L; Molecular Biotechnology, Department of Biosciences, Goethe University Frankfurt, 60438, Frankfurt am Main, Germany.; State Key Laboratory of Bio-organic and Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai, 200032, China., Bode HB; Department of Natural Products in Organismic Interactions, Max-Planck Institute for Terrestrial Microbiology, 35043, Marburg, Germany. helge.bode@mpi-marburg.mpg.de.; Molecular Biotechnology, Department of Biosciences, Goethe University Frankfurt, 60438, Frankfurt am Main, Germany. helge.bode@mpi-marburg.mpg.de.; Department of Chemistry, Chemical Biology, Phillips University Marburg, 35043, Marburg, Germany. helge.bode@mpi-marburg.mpg.de.; Center for Synthetic Microbiology (SYNMIKRO), Phillips University Marburg, 35043, Marburg, Germany. helge.bode@mpi-marburg.mpg.de.; Senckenberg Gesellschaft für Naturforschung, 60325, Frankfurt, Germany. helge.bode@mpi-marburg.mpg.de.
المصدر: Microbial cell factories [Microb Cell Fact] 2024 Apr 01; Vol. 23 (1), pp. 98. Date of Electronic Publication: 2024 Apr 01.
نوع المنشور: Journal Article
اللغة: English
بيانات الدورية: Publisher: BioMed Central Country of Publication: England NLM ID: 101139812 Publication Model: Electronic Cited Medium: Internet ISSN: 1475-2859 (Electronic) Linking ISSN: 14752859 NLM ISO Abbreviation: Microb Cell Fact Subsets: MEDLINE
أسماء مطبوعة: Original Publication: London : BioMed Central, [2002-
مواضيع طبية MeSH: Xenorhabdus*/genetics , Xenorhabdus*/metabolism , Photorhabdus*/genetics , Biological Products*/metabolism, Gene Editing ; Clustered Regularly Interspaced Short Palindromic Repeats
مستخلص: Background: Bacteria of the genus Photorhabdus and Xenorhabdus are motile, Gram-negative bacteria that live in symbiosis with entomopathogenic nematodes. Due to their complex life cycle, they produce a large number of specialized metabolites (natural products) encoded in biosynthetic gene clusters (BGC). Genetic tools for Photorhabdus and Xenorhabdus have been rare and applicable to only a few strains. In the past, several tools have been developed for the activation of BGCs and the deletion of individual genes. However, these often have limited efficiency or are time consuming. Among the limitations, it is essential to have versatile expression systems and genome editing tools that could facilitate the practical work.
Results: In the present study, we developed several expression vectors and a CRISPR-Cpf1 genome editing vector for genetic manipulations in Photorhabdus and Xenorhabdus using SEVA plasmids. The SEVA collection is based on modular vectors that allow exchangeability of different elements (e.g. origin of replication and antibiotic selection markers with the ability to insert desired sequences for different end applications). Initially, we tested different SEVA vectors containing the broad host range origins and three different resistance genes for kanamycin, gentamycin and chloramphenicol, respectively. We demonstrated that these vectors are replicative not only in well-known representatives, e.g. Photorhabdus laumondii TTO1, but also in other rarely described strains like Xenorhabdus sp. TS4. For our CRISPR/Cpf1-based system, we used the pSEVA231 backbone to delete not only small genes but also large parts of BGCs. Furthermore, we were able to activate and refactor BGCs to obtain high production titers of high value compounds such as safracin B, a semisynthetic precursor for the anti-cancer drug ET-743.
Conclusions: The results of this study provide new inducible expression vectors and a CRISPR/CPf1 encoding vector all based on the SEVA (Standard European Vector Architecture) collection, which can improve genetic manipulation and genome editing processes in Photorhabdus and Xenorhabdus.
(© 2024. The Author(s).)
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فهرسة مساهمة: Keywords: Photorhabdus; Xenorhabdus; CRISPR/Cpf1; Genome editing; Natural products; Safracin
المشرفين على المادة: 0 (Biological Products)
تواريخ الأحداث: Date Created: 20240402 Date Completed: 20240403 Latest Revision: 20240404
رمز التحديث: 20240404
مُعرف محوري في PubMed: PMC10983751
DOI: 10.1186/s12934-024-02363-8
PMID: 38561780
قاعدة البيانات: MEDLINE
الوصف
تدمد:1475-2859
DOI:10.1186/s12934-024-02363-8