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Full Description
This volume details methods ranging from environmental isolation, small-scale to large-scale cell cultivation, genome sequencing, genome editing, and "omics" studies. Chapters provide reliable and practical resources that strengthen the collective toolkit of the U. maydis community and fosters further discoveries in fungal biology, plant pathology, applied microbiology and beyond. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips for troubleshooting and avoiding known pitfalls.
Authoritative and up-to-date, Ustilago maydis: Methods and Protocols aims to be comprehensive guide for researchers in the field
Contents
Induction medium to analyze in vitro morphological changes and biotrophic features in sporidia of Ustilago maydis.- Induction of Ustilago maydis appressorium and filamentation.- Ustilago maydis Seedling Infection assay and scoring.- Apoplastic Fluid Collection/Analysis of Ustilago maydis-Infected Maize Leaves.- Chlorazol Black E staining of plant tissue infected with Ustilago maydis to monitor pathogenic development on and in planta.- Wheat germ agglutinin-Alexa Fuor488/propidium iodine co-staining and fluorescence microscopic imaging of Ustilago maydis infected maize anthers.- A genomic DNA-based real-time PCR assay for quantification of Ustilago maydis biomass in plant samples.- Random (UV) mutagenesis, unbiased screening and identification of DNA repair mutants by complementation cloning.- Synchronous induction and microscopic analysis of hyphal growth using laboratory strain AB33.- maydis Protoplast Generation and Transformation.- Isolation of genomic DNA from Ustilago species.- An efficient colony PCR method for construct verification in the basidiomycete Ustilago maydis.- Southern blotting: A valuable molecular technique to verify gene replacement in Ustilago maydis.- Golden Gate assembly for genome editing in Ustilago maydis.- Generation of gene knock-out mutants in Ustilago maydis using Cas9hf nuclease.- FLP/FRT-mediated recombination for resistance marker recycling.- Conditional gene deletion in Ustilago maydis via CRE recombinase in planta.- Efficient introduction of genetic elements at specific insertion loci.- Viability of Ustilago maydis by Flow Cytometry.- Evaluating Gene Expression in Ustilago maydis using RT-qPCR.- Isolation of RNA for RNA sequencing in Ustilago maydis.- Determination of spontaneous mutation rate by Fluctuation Analysis using 5-Fluoroorotic acid (5-FOA).- Insertion Pool Sequencing (iPool-Seq) for the high-throughput quantification of Ustilago maydis mutant fitness.- Mitotic heteroallelic and sister chromatid recombination.- The Use of Yeast Two-Hybrid Analysis to Study Protein-Protein Interactions for Ustilago maydis.- TurboID-based proximity labeling for exploiting effector interacting proteins in the fungal maize pathogen Ustilago maydis.- Immunodetection of Cell-Surface-Associated Proteins in Ustilago maydis and Infected Maize Leaves.- Protein Extraction Methods for Ustilago maydis.- Studying cell wall-associated effectors in U. maydis.- Computational Identification and Structural Analysis of Ustilago maydis Effector Proteins.- Chromatin Immunoprecipitation (ChIP) to investigate histone-modifications in smut fungi.- Expression and detection of heterologous proteins in Ustilago maydis.- Mannosylerythritol lipid and ustilagic acid analysis using thin-layer chromatography.- Studying Peroxisomes in Ustilago maydis: A Defined Workflow .- Methods for studying the polyamines in Ustilago maydis.- Extraction of total fatty acids from Ustilago maydis.- Metabolomic study in Ustilaginaceae with GC-MS/MS.- ER and cell wall stress assay of Ustilago maydis strains.- Simultaneous Saccharification and Fermentation with Ustilago maydis and Analytical Methods for Cell and Product Quantification.- BN-PAGE of Ustilago maydis respiratory complexes.- Trehalose Determination in Ustilago maydis under Stress Conditions .- H2O2 consumption and production assay for the characterization of U. maydis oxidoreductases.- Online monitoring of oxygen transfer rate as a versatile tool for fermentation characterization and optimization.- Online monitoring of U. maydis utilizing fluorescence and scattered light measurements in microtiter plate cultivations.- Integrating pH value and base consumption in online monitoring of organic acid production.- Online monitor the viable biomass formation of U. maydis by capacitance measurement in lab scale fermenter.
