detection of polymyxa betae in sugar beet roots using rt-pcr method in razavi khorasan province

abstract polymyxa betae is an obligate parasite and has a worldwide distribution. this fungus is a soil-borne root endoparasite in some plant families such as portulaceae, chenopodiaceae and amaranthaceae. this fungus is the vector of many plant viruses such as beet necrotic yellow vein virus (bnyvv) which causes the disease known as rhizomania. bnyvv is the most economical important viral diseases of sugar beet. resting spores of p.betae can survive in soil for many years. for detection of this fungus in razavi khorasan province, in summer of 2005, soil samples and plants with rhizomania symptoms were collected from the sugar beet fields. infection of samples with bnyvv was confirmed by das-elisa. baiting technique was used for isolation of the fungus. six-week old plant roots were taken from soil, washed and stained with koh 10% for 30 minutes and roots were examined for the presence of resting spores under light microscope. due to difficulty in observing the immature resting spores of the fungus and distinction between zoosporangium and resting spores of other fungi by light microscope, pcr technique was used to the detection of the fungus. total rna was extracted from roots of infected samples using peg6000 precipitation method and cdna was made using fungus specific forward primer. pcr was performed with the specific forward and reverse primers. after electrophoresis on 1.5% agarose gel and 5% polyacrylamid gel, the band of 170 bp was detected. this amplified region is specific for p.betae.