title: An efficient method for the extraction of high-quality fungal total RNA to study the Mycosphaerella fijiensis-Musa spp. Interaction.
abstract: Efficient RNA isolation is a prerequisite for gene expression studies and it has an increasingly important role in the study of plant-fungal pathogen interactions. However, RNA isolation is difficult in filamentous fungi. These organisms are notorious for their rigid cell walls and the presence of high levels of carbohydrates, excreted from the fungal cells during submerged growth, which interferes with the extraction procedures. Although many commercial kits are already available for RNA isolation, they do not provide, in most cases, enough amount of pure RNA to be used in upstream applications. In the present work, we propose an easy and efficient protocol for isolating total RNA from the filamentous fungus Mycosphaerella fijiensis, the most important foliar pathogen of Musa spp. varieties worldwide. In addition, we applied the proposed protocol to the isolation of total RNA from banana leaves infected with the pathogen. Our methodology was developed based on the SDS method with modifications including a carbohydrate precipitation step. The protocol resulted in high-quality total RNA, from fungal mycelium grown in PDB medium and infected banana leaves, suitable for further molecular studies. The proposed methodology is also applicable to the ascomycete fungus Passalora fulva (syn. Cladosporum fulvum).
authors.affiliation: Instituto de Biotecnología de las Plantas (IBP), Universidad Central Marta Abreu de Las Villas, Carretera a Camajuaní km 5.5, 54830 Santa Clara, Cuba.
authors.forename: Aminael
authors.initials: A
authors.lastname: Sánchez-Rodríguez
authors.forename: Orelvis
authors.initials: O
authors.lastname: Portal
authors.forename: Luis E
authors.initials: LE
authors.lastname: Rojas
authors.forename: Bárbara
authors.initials: B
authors.lastname: Ocaña
authors.forename: Milady
authors.initials: M
authors.lastname: Mendoza
authors.forename: Mayra
authors.initials: M
authors.lastname: Acosta
authors.forename: Elio
authors.initials: E
authors.lastname: Jiménez
authors.forename: Monica
authors.initials: M
authors.lastname: Höfte
mesh_terms: D001203:Ascomycota / Q000235:genetics*; D054884:Host-Pathogen Interactions; D008967:Molecular Biology / Q000379:methods*; D028521:Musa / Q000382:microbiology*; D025282:Mycelium; D010935:Plant Diseases / Q000382:microbiology; D018515:Plant Leaves / Q000382:microbiology; D012331:RNA, Fungal / Q000302:isolation & purification* / Q000592:standards*
chemical_list: D012331:RNA, Fungal