A non-phenol-chloroform extraction of double-stranded RNA from plant and fungal tissues.
Double-stranded RNA (dsRNA) molecules of viruses are found in nature at a very high frequency. Their detection in plants and fungi has been carried out with difficulty due to the complicated dsRNA extraction techniques used commonly which includes phenol-chloroform extractions. In this study, an extraction method for isolation of dsRNA is described that is free of phenol and chloroform. A lysis buffer, containing beta-mercaptoethanol and polyvinylpolypyrrolidone (PVPP-40), was added to homogenised tissues and the subsequent supernatant was filtered through a cellulose CF-11 mini-column. DsRNA molecules were separated based on the differing affinity of nucleic acids for the cellulose CF-11 resin in 20% ethanol buffer. This easy, rapid and cheap technique has been successfully tested on fungi and plants containing different dsRNA virus molecules, indicating the possibility of a wide use of the method.
Balijja
Alitukiriza
A
CNR, Istituto per la Protezione delle Piante, 50019 Sesto Fiorentino, Via Madonna del Piano 10, Italy. balijja@ipp.cnr.it
Kvarnheden
Anders
A
Turchetti
Tullio
T
D004271:DNA, Fungal / Q000302:isolation & purification; D018744:DNA, Plant / Q000302:isolation & purification; D005658:Fungi / Q000821:virology*; D012328:RNA Viruses / Q000235:genetics*; D012330:RNA, Double-Stranded / Q000302:isolation & purification*; D012367:RNA, Viral / Q000302:isolation & purification*; D014026:Nicotiana / Q000821:virology
D004271:DNA, Fungal; D018744:DNA, Plant; D012330:RNA, Double-Stranded; D012367:RNA, Viral