An improved and reproducible protocol for the extraction of high quality fungal RNA from plant biomass substrates

• Published in: Fungal genetics and biology 2014-11, Vol.72, p.201-206
• Main Authors: Patyshakuliyeva, Aleksandrina, Mäkelä, Miia R., Sietiö, Outi-Maaria, de Vries, Ronald P., Hildén, Kristiina S.
• Format: Article
• Language: English
• Subjects: Agaricus bisporus; Biomass; Centrifugation, Density Gradient - methods; CsCl gradient ultracentrifugation; Dichomitus squalens; Gene expression; Gene Expression Profiling - methods; Lignocellulose (plant biomass); Molecular Biology - methods; Plants - microbiology; RNA extraction; RNA, Fungal - isolation & purification
• ISSN: 1087-1845; 1096-0937
• DOI: 10.1016/j.fgb.2014.06.001

•A method was developed to isolate high quality RNA from plant biomass cultures.•The extraction was highly reproducible.•The methods allowed extraction even from compost grown fungi.•The method resulted in high quality and quantity RNA which was very suitable for Q-PCR and transcriptomics. Isolation of high quantity and quality RNA is a crucial step in the detection of meaningful gene expression data. Obtaining intact fungal RNA from complex lignocellulosic substrates is often difficult, producing low integrity RNA which perform poorly in downstream applications. In this study we developed an RNA extraction method using CsCl centrifugation procedure, modified from previous reports and adapted for isolation of RNA from plant biomass. This method provided high level of integrity and good quantity of RNA which were suitable for reliable analyses of gene expression and produced consistent and reproducible results.