Genomic Edition of Ashbya gossypii Using One-vector CRISPR/Cas9

The CRISPR/Cas9 system is a novel genetic tool which allows the precise manipulation of virtually any genomic sequence. In this protocol, we use a specific CRISPR/Cas9 system for the manipulation of Ashbya gossypii. The filamentous fungus A. gossypii is currently used for the industrial production of riboflavin (vitamina B2). In addition, A. gossypii produces other high-value compounds such as folic acid, nucleosides and biolipids. A large molecular toolbox is available for the genomic manipulation of this fungus including gene targeting methods, rapid assembly of heterologous expression modules and, recently, a one-vector CRISPR/Cas9 editing system adapted for A. gossypii that allows marker-free engineering strategies to be implemented. The CRISPR/Cas9 system comprises an RNA guided DNA endonuclease (Cas9) and a guide RNA (gRNA), which is complementary to the genomic target region. The Cas9 nuclease requires a 5′-NGG-3′ trinucleotide, called protospacer adjacent motif (PAM), to generate a double-strand break (DSB) in the genomic target, which can be repaired with a synthetic mutagenic donor DNA (dDNA) by homologous recombination (HR), thus introducing a specific designed mutation. The CRISPR/Cas9 system adapted for A. gossypii largely facilitates the genomic edition of this industrial fungus

Analysis of Undergraduate Dermatology Syllabi at Spanish Universities: Does the Weight of Theoretical Content Match the Skin Conditions Seen in Primary Care and General Dermatology Practices?

Introduction: Undergraduate dermatology courses vary in the nearly 50 Spanish medical faculties that teach the subject. This study aimed to describe the characteristics of these courses and to analyze whether the weight assigned to dermatology topics reflects the caseloads of primary care physicians and general dermatologists in the Spanish national health system. Material and methods: Cross-sectional study of syllabi used in Spanish medical faculties during the 2021---2022 academic year. We determined the number of teaching hours in public and private university curricula and compared the weight of dermatology topics covered to the dermatology caseloads of primary care physicians and general dermatologists as reported in published studies. Results: Most medical faculties taught dermatology for one semester. The median number of credits offered was 4.5. On average, lectures covered 24 theoretical topics, and seminars and workshops covered 9 topics. We identified a clear disparity between the percentage of time devoted to dermatology topics in course lectures and the skin conditions usually managed in primary care and general dermatology practices

New Promoters for Metabolic Engineering of Ashbya gossypii

[EN]Ashbya gossypii is a filamentous fungus that is currently exploited for the industrial production of riboflavin. In addition, metabolically engineered strains of A. gossypii have also been described as valuable biocatalysts for the production of different metabolites such as folic acid, nucleosides, and biolipids. Hence, bioproduction in A. gossypii relies on the availability of well-performing gene expression systems both for endogenous and heterologous genes. In this regard, the identification of novel promoters, which are critical elements for gene expression, decisively helps to expand the A. gossypii molecular toolbox. In this work, we present an adaptation of the Dual Luciferase Reporter (DLR) Assay for promoter analysis in A. gossypii using integrative cassettes. We demonstrate the efficiency of the analysis through the identification of 10 new promoters with different features, including carbon source-regulatable abilities, that will highly improve the gene expression platforms used in A. gossypii. Three novel strong promoters (PCCW12, PSED1, and PTSA1) and seven medium/weak promoters (PHSP26, PAGL366C, PTMA10, PCWP1, PAFR038W, PPFS1, and PCDA2) are presented. The functionality of the promoters was further evaluated both for the overexpression and for the underexpression of the A. gossypii MSN2 gene, which induced significant changes in the sporulation ability of the mutant strains