genome editing
Commentary | Molecular Biology and PhysiologymSphere of Influence: Ushering in the CRISPR Revolution to Toxoplasma BiologyAlfredo J. Guerra works in the field of molecular parasitology and structural biology. In this mSphere of Influence article, he reflects on how “Efficient Gene Disruption in Diverse Strains of Toxoplasma gondii Using CRISPR/CAS9” by Bang Shen et al. (mBio 5:e01114-14, 2014, https://doi.org/10.1128/mBio.01114-14) and...
Research Article | Synthetic BiologyDevelopment of a CRISPR/Cas9-Based Tool for Gene Deletion in Issatchenkia orientalisMicrobial production of fuels and chemicals from renewable and readily available biomass is a sustainable and economically attractive alternative to petroleum-based production. Because of its unusual tolerance to highly acidic conditions, I. orientalis is a promising potential candidate for the manufacture of valued organic acids. Nevertheless, reliable and efficient genetic engineering tools in I. orientalis are...
Research Article | Host-Microbe BiologyOptimizing Systems for Cas9 Expression in Toxoplasma gondiiToxoplasma gondii is an intracellular parasite that causes life-threatening disease in immunocompromised patients and affects the developing fetus when contracted during pregnancy. Closely related species cause malaria and severe diarrhea, thereby constituting leading causes for childhood mortality. Despite their importance to global health, this family of parasites...
Research Article | Molecular Biology and PhysiologyGenome Editing of Babesia bovis Using the CRISPR/Cas9 SystemBabesia bovis is the most virulent cause of bovine babesiosis worldwide. The disease consequences are death, abortion, and economical loss due to reduced milk and meat production. Available vaccines are not effective, treatment options are limited, and emergence of drug and acaricide resistance has been reported from different regions. There is an urgent need to...
Editor's Pick Research Article | Molecular Biology and PhysiologyPlasmid-Based CRISPR-Cas9 Gene Editing in Multiple Candida SpeciesCandida species are a major cause of infection worldwide. The species associated with infection vary with geographical location and with patient population. Infection with Candida tropicalis is particularly common in South America and Asia, and Candida parapsilosis infections are more...
Editor's Pick Research Article | Molecular Biology and PhysiologyA Simple and Universal System for Gene Manipulation in Aspergillus fumigatus: In Vitro-Assembled Cas9-Guide RNA Ribonucleoproteins Coupled with Microhomology Repair TemplatesTackling the multifactorial nature of virulence and antifungal drug resistance in A. fumigatus requires the mechanistic interrogation of a multitude of genes, sometimes across multiple genetic backgrounds. Classical fungal gene replacement systems can be laborious and time-consuming and, in wild-type isolates, are impeded by low rates of homologous recombination. Our...
Research Article | Host-Microbe BiologyUse of RNA-Protein Complexes for Genome Editing in Non-albicans Candida SpeciesExisting CRISPR-Cas9 genome modification systems for use in Candida albicans, which rely on constructs to endogenously express the Cas9 protein and guide RNA, do not work efficiently in other Candida species due to inefficient promoter activity. Here, we present an expression-free method that uses RNA-protein complexes and demonstrate its use in three ...
Editor's Pick Research Article | Molecular Biology and PhysiologyAn Efficient, Rapid, and Recyclable System for CRISPR-Mediated Genome Editing in Candida albicansCandida albicans is the major fungal pathogen of humans and is the subject of intense biomedical and discovery research. Until recently, the pace of research in this field has been hampered by the lack of efficient methods for genome editing. We report the development of a highly efficient and flexible genome editing system for use with...
Research Article | Host-Microbe BiologyElectric Cell-Substrate Impedance Sensing To Monitor Viral Growth and Study Cellular Responses to Infection with Alphaherpesviruses in Real TimeAlphaherpesviruses, including those that commonly infect humans, such as HSV-1 and HSV-2, typically infect and cause cellular damage to epithelial cells at mucosal surfaces, leading to disease. The development of novel technologies to study the cellular responses to infection may allow a more complete understanding of virus replication and the creation of novel antiviral therapies. This study demonstrates the use of ECIS to study...
Research Article | Molecular Biology and PhysiologyOptimized CRISPR-Cas9 Genome Editing for Leishmania and Its Use To Target a Multigene Family, Induce Chromosomal Translocation, and Study DNA Break Repair MechanismsLeishmania parasites cause human leishmaniasis. To accelerate characterization of Leishmania genes for new drug and vaccine development, we optimized and simplified the CRISPR-Cas9 genome-editing tool for Leishmania. We show that co-CRISPR targeting of the miltefosine transporter gene and serial transfections of an oligonucleotide donor significantly eased isolation of edited mutants. This cotargeting strategy...