دورية أكاديمية
أعرض في EDS

Development of a telomere vector-based approach to overcome limitations caused by lethal phenotypes in the study of essential genes in Magnaporthe oryzae.

التفاصيل البيبلوغرافية
العنوان: Development of a telomere vector-based approach to overcome limitations caused by lethal phenotypes in the study of essential genes in Magnaporthe oryzae.
المؤلفون: Wirtz L; Department of Molecular Plant Physiology, RWTH Aachen University, Aachen, Germany., Casanova F; Department of Molecular Plant Physiology, RWTH Aachen University, Aachen, Germany., Schaffrath U; Department of Molecular Plant Physiology, RWTH Aachen University, Aachen, Germany., Wegner A; Department of Molecular Plant Physiology, RWTH Aachen University, Aachen, Germany.
المصدر: Molecular plant pathology [Mol Plant Pathol] 2024 May; Vol. 25 (5), pp. e13460.
نوع المنشور: Journal Article; Research Support, Non-U.S. Gov't
اللغة: English
بيانات الدورية: Publisher: Blackwell Science in collaboration with the British Society of Plant Pathology Country of Publication: England NLM ID: 100954969 Publication Model: Print Cited Medium: Internet ISSN: 1364-3703 (Electronic) Linking ISSN: 13643703 NLM ISO Abbreviation: Mol Plant Pathol Subsets: MEDLINE
أسماء مطبوعة: Original Publication: Oxford, UK : Blackwell Science in collaboration with the British Society of Plant Pathology, c2000-
مواضيع طبية MeSH: Telomere*/genetics , Phenotype* , Genes, Essential* , Genetic Vectors*/genetics , Ascomycota*, CRISPR-Cas Systems/genetics ; Genes, Fungal/genetics ; Gene Deletion ; Magnaporthe/genetics ; Magnaporthe/pathogenicity
مستخلص: Reverse genetic approaches are common tools in genomics for elucidating gene functions, involving techniques such as gene deletion followed by screening for aberrant phenotypes. If the generation of gene deletion mutants fails, the question arises whether the failure stems from technical issues or because the gene of interest (GOI) is essential, meaning that the deletion causes lethality. In this report, we introduce a novel method for assessing gene essentiality using the phytopathogenic ascomycete Magnaporthe oryzae. The method is based on the observation that telomere vectors are lost in transformants during cultivation without selection pressure. We tested the hypothesis that essential genes can be identified in deletion mutants co-transformed with a telomere vector. The M. oryzae gene MoPKC, described in literature as essential, was chosen as GOI. Using CRISPR/Cas9 technology transformants with deleted GOI were generated and backed up by a telomere vector carrying a copy of the GOI and conferring fenhexamid resistance. Transformants in which the GOI deletion in the genome was not successful lost the telomere vector on media without fenhexamid. In contrast, transformants with confirmed GOI deletion retained the telomere vector even in absence of fenhexamid selection. In the latter case, the maintenance of the telomere indicates that the GOI is essential for the surveillance of the fungi, as it would have been lost otherwise. The method presented here allows to test for essentiality of genes when no mutants can be obtained from gene deletion approaches, thereby expanding the toolbox for studying gene function in ascomycetes.
(© 2024 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)
References: Pharmacol Ther. 2002 Jul;95(1):1-20. (PMID: 12163125)
Genetics. 1995 Jun;140(2):479-92. (PMID: 7498730)
Mol Microbiol. 2015 Oct;98(3):403-19. (PMID: 26192090)
Nucleic Acids Res. 1997 Apr 1;25(7):1426-31. (PMID: 9060439)
Gene. 1990 Apr 16;88(2):159-65. (PMID: 1971801)
Nat Commun. 2015 Mar 31;6:6741. (PMID: 25824284)
Genetics. 2002 Jul;161(3):1077-87. (PMID: 12136012)
Philos Trans R Soc Lond B Biol Sci. 2016 Dec 5;371(1709):. (PMID: 28080995)
Gene. 1993 Sep 30;132(1):67-73. (PMID: 8104848)
Gene. 1992 Aug 1;117(1):81-9. (PMID: 1644317)
Mol Cell Biol. 1987 May;7(5):1725-30. (PMID: 3600642)
Fungal Genet Biol. 1998 Oct;25(1):22-30. (PMID: 9806803)
Eukaryot Cell. 2011 Sep;10(9):1148-55. (PMID: 21724934)
PLoS Pathog. 2020 Aug 17;16(8):e1008326. (PMID: 32804988)
Genome Res. 2003 Mar;13(3):399-406. (PMID: 12618370)
Biosci Biotechnol Biochem. 2007 Nov;71(11):2787-99. (PMID: 17986778)
Plant Cell. 1993 Nov;5(11):1575-90. (PMID: 8312740)
Mol Microbiol. 1992 Dec;6(23):3603-10. (PMID: 1474902)
Sci Rep. 2018 Sep 25;8(1):14355. (PMID: 30254203)
Mol Microbiol. 2012 Feb;83(3):506-19. (PMID: 22220813)
Fungal Genet Biol. 2008 Jan;45(1):68-75. (PMID: 17716934)
Eukaryot Cell. 2003 Apr;2(2):247-55. (PMID: 12684374)
Eur Respir J Suppl. 2002 Jul;36:78s-86s. (PMID: 12168750)
Eukaryot Cell. 2005 Feb;4(2):487-9. (PMID: 15701811)
Mol Microbiol. 1999 Nov;34(4):792-8. (PMID: 10564518)
Nucleic Acids Res. 1993 Feb 11;21(3):497-504. (PMID: 8441663)
Trends Microbiol. 2016 Jan;24(1):35-50. (PMID: 26549580)
Mol Microbiol. 2003 Oct;50(1):167-81. (PMID: 14507372)
Science. 1999 Aug 6;285(5429):901-6. (PMID: 10436161)
Mol Plant Pathol. 2024 May;25(5):e13460. (PMID: 38695626)
J Bacteriol. 1990 Jun;172(6):3163-71. (PMID: 2345140)
معلومات مُعتمدة: RWTH Aachen University scholarships for Doctoral Students; SCHA 631/11-1 Deutsche Forschungsgemeinschaft
فهرسة مساهمة: Keywords: Magnaporthe oryzae; CRISPR/Cas9; gene deletion; mutational analyses; reverse genetics; rice blast
SCR Organism: Pyricularia oryzae
تواريخ الأحداث: Date Created: 20240502 Date Completed: 20240502 Latest Revision: 20240504
رمز التحديث: 20240504
مُعرف محوري في PubMed: PMC11064798
DOI: 10.1111/mpp.13460
PMID: 38695626
قاعدة البيانات: MEDLINE
الوصف
تدمد:1364-3703
DOI:10.1111/mpp.13460