テンサイシストセンチュウ汚染土壌からの迅速なdna検出方法

Abstract

Immediately after the infestation was identified, the fields were treated with pesticides, including fumigants, to exterminate the nematodes. However, cysts can survive these chemical treatments, and nematode populations in these fields can recover, especially with renewed cultivation where the vegetables become hosts for the nematodes. It is important to carefully monitor these fields for the re-establishment of nematode populations. Rapid nematode detection methods are therefore needed.Numerous methods of identifying cyst nematodes, based on DNA extraction followed by PCR, have been described. These methods involve the separation of nematodes, either as cysts or juveniles, from the soil, followed by direct DNA extraction from nematode bodies. Specific primers have been developed for species identification. PCR-RFLP of ITS-rDNA can distinguish between the four Heterodera species, H. schachtii, H. betae, H. trifolii, and H. medicaginis (Amiri et al., 2002). DNA has also been extracted from soil samples. Species-specific primers were used in combination with realtime PCR to quantify H. glycines in 30 g samples of dried soil pulverized with a ball mill (Shirai and Toyota, 2019).Methods of extracting DNA from soil samples and identifying eukaryotic organisms in these samples include environmental DNA sequencing, also called metabarcoding. Soil samples are immersed in saturated phosphate buffer (0.12 M Na 2 HPO 4 /NaH 2 PO 4 , pH 8.0) to release DNA bound to soil