دورية أكاديمية
أعرض في EDS

Bidirectional titration of yeast gene expression using a pooled CRISPR guide RNA approach.

التفاصيل البيبلوغرافية
العنوان: Bidirectional titration of yeast gene expression using a pooled CRISPR guide RNA approach.
المؤلفون: Bowman EK; Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, TX, 78712., Deaner M; McKetta Department of Chemical Engineering, The University of Texas at Austin, Austin, TX, 78712., Cheng JF; The US Department of Energy Joint Genome Institute, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720., Evans R; The US Department of Energy Joint Genome Institute, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720., Oberortner E; The US Department of Energy Joint Genome Institute, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720., Yoshikuni Y; The US Department of Energy Joint Genome Institute, Lawrence Berkeley National Laboratory, Berkeley, CA, 94720., Alper HS; Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin, TX, 78712; halper@che.utexas.edu.; McKetta Department of Chemical Engineering, The University of Texas at Austin, Austin, TX, 78712.
المصدر: Proceedings of the National Academy of Sciences of the United States of America [Proc Natl Acad Sci U S A] 2020 Aug 04; Vol. 117 (31), pp. 18424-18430. Date of Electronic Publication: 2020 Jul 20.
نوع المنشور: Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.
اللغة: English
بيانات الدورية: Publisher: National Academy of Sciences Country of Publication: United States NLM ID: 7505876 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1091-6490 (Electronic) Linking ISSN: 00278424 NLM ISO Abbreviation: Proc Natl Acad Sci U S A Subsets: MEDLINE
أسماء مطبوعة: Original Publication: Washington, DC : National Academy of Sciences
مواضيع طبية MeSH: RNA, Fungal/*genetics , RNA, Guide, CRISPR-Cas Systems/*genetics , Saccharomyces cerevisiae/*genetics, CRISPR-Cas Systems ; Clustered Regularly Interspaced Short Palindromic Repeats ; Gene Expression Regulation, Fungal ; Gene Library ; Plasmids/genetics ; Plasmids/metabolism ; Promoter Regions, Genetic ; RNA, Fungal/metabolism ; RNA, Guide, CRISPR-Cas Systems/metabolism ; Saccharomyces cerevisiae/metabolism
مستخلص: Most classic genetic approaches utilize binary modifications that preclude the identification of key knockdowns for essential genes or other targets that only require moderate modulation. As a complementary approach to these classic genetic methods, we describe a plasmid-based library methodology that affords bidirectional, graded modulation of gene expression enabled by tiling the promoter regions of all 969 genes that comprise the ito977 model of Saccharomyces cerevisiae' s metabolic network. When coupled with a CRISPR-dCas9-based modulation and next-generation sequencing, this method affords a library-based, bidirection titration of gene expression across all major metabolic genes. We utilized this approach in two case studies: growth enrichment on alternative sugars, glycerol and galactose, and chemical overproduction of betaxanthins, leading to the identification of unique gene targets. In particular, we identify essential genes and other targets that were missed by classic genetic approaches.
Competing Interests: The authors declare no competing interest.
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فهرسة مساهمة: Keywords: CRISPR; essential genes; graded expression; next-generation sequencing; synthetic biology
المشرفين على المادة: 0 (RNA, Fungal)
0 (RNA, Guide, CRISPR-Cas Systems)
تواريخ الأحداث: Date Created: 20200722 Date Completed: 20200930 Latest Revision: 20240104
رمز التحديث: 20240104
مُعرف محوري في PubMed: PMC7414176
DOI: 10.1073/pnas.2007413117
PMID: 32690674
قاعدة البيانات: MEDLINE
الوصف
تدمد:1091-6490
DOI:10.1073/pnas.2007413117