A RAPID AND EFFECTIVE METHOD FOR TOTAL RNA EXTRACTION FROM CONIDIA AND MYCELIUM OF SEPTORIA TRITICI

Abstract

Abstract An effective and rapid method for RNA extraction from conidia and mycelium of S. tritici was developed, in which guanidinium isothiocyanate, CTAB and insoluble PVP were combined in one step, simplifying the extraction process greatly. The procedure can be completed in less than 6 h. Yield and purity of RNA could be estimated by electrophoresis on 1% formaldhyde-denatured agarose gel and by measuring the absorbance at 280, 260, 230 and 210 nm by UV spectroscopy. The result of electrophoresis showed two clear bands of 18S and 28S rRNA and no DNA contamination in the isolated RNA solution. The ratios A260/A280, A260/A230, A260/A210 were 1.76–1.82, 2.79–3.21 and 0.81–0.86, respectively. A high yield with 532–881 ng total RNA per gram fresh conidia and mycelium of S. tritici was obtained. The RNA was of sufficient purity for RT-PCR to detect the expression of β-tubulin gene in S. tritici. This protocol may also be suitable for RNA extraction from spores and mycelium of other fungi.