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dc.contributor.authorAfonina, Irinaen_US
dc.contributor.authorOng, Juneen_US
dc.contributor.authorChua, Jeromeen_US
dc.contributor.authorLu, Timothyen_US
dc.contributor.authorKline, Kimberly A.en_US
dc.identifier.citationAfonina, I., Ong, J., Chua, J., Lu, T., & Kline, K. A. Multiplex CRISPRi system enables the study of stage-specific biofilm genetic requirements in Enterococcus faecalis. mBio, 11(5), e01101-20-. doi:10.1128/mBio.01101-20en_US
dc.description.abstractEnterococcus faecalis is an opportunistic pathogen, which can cause multidrug-resistant life-threatening infections. Gaining a complete understanding of enterococcal pathogenesis is a crucial step in identifying a strategy to effectively treat enterococcal infections. However, bacterial pathogenesis is a complex process often involving a combination of genes and multilevel regulation. Compared to established knockout methodologies, CRISPR interference (CRISPRi) approaches enable the rapid and efficient silencing of genes to interrogate gene products and pathways involved in pathogenesis. As opposed to traditional gene inactivation approaches, CRISPRi can also be quickly repurposed for multiplexing or used to study essential genes. Here, we have developed a novel dual-vector nisin-inducible CRISPRi system in E. faecalis that can efficiently silence via both nontemplate and template strand targeting. Since the nisin-controlled gene expression system is functional in various Gram-positive bacteria, the developed CRISPRi tool can be extended to other genera. This system can be applied to study essential genes, genes involved in antimicrobial resistance, and genes involved in biofilm formation and persistence. The system is robust and can be scaled up for high-throughput screens or combinatorial targeting. This tool substantially enhances our ability to study enterococcal biology and pathogenesis, host-bacterium interactions, and interspecies communication.en_US
dc.description.sponsorshipMinistry of Education (MOE)en_US
dc.description.sponsorshipNational Research Foundation (NRF)en_US
dc.rights© 2020 Afonina et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license.en_US
dc.subjectScience::Biological sciencesen_US
dc.titleMultiplex CRISPRi system enables the study of stage-specific biofilm genetic requirements in Enterococcus faecalisen_US
dc.typeJournal Articleen
dc.contributor.schoolSchool of Biological Sciencesen_US
dc.contributor.researchSingapore Centre for Environmental Life Sciences and Engineering (SCELSE)en_US
dc.description.versionPublished versionen_US
dc.subject.keywordsEnterococcus Faecalisen_US
dc.subject.keywordsCRISPR Interferenceen_US
dc.description.acknowledgementThis research is supported by the National Research Foundation, Prime Minister’sOffice, Singapore, under its Campus for Research Excellence and Technological Enterprise (CREATE) program. The work was carried out through core funding of the Singapore-MIT Alliance for Research and Technology (SMART) Antimicrobial Resistance Interdisciplinary Research Group (AMR IRG). Part of the work was carried out at the Singapore Centre for Environmental and Life Science Engineering (SCELSE), whose research is supported by the National Research Foundation Singapore, Ministry of Education, to Nanyang Technological University and the National University of Singapore under its Research Centre of Excellence Programme.en_US
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