Establishment of the Loop-mediated Isothermal Amplification Assay for Detection of Phytophthora parasitica

In order to establish a rapid detection method for Phytophthora parasitica, hydroxynaphthol blue (HNB) was used as an indicator for the Phytophthora nicotiana specific transcribed spacer (ITS) DNA sequence. The primers were designed with the LAMP design software. The LAMP reaction system and reaction conditions were optimized and the specificity, sensitivity and practical application of the detection method were verified. The results showed that the LAMP reaction system of Phytophthora nicotianae established in this experiment has good specificity and high sensitivity. Specific detection showed that only the LAMP product of the Phytophthora nicotiana strain is positive (blue), and the electrophoresis can produce trapezoidal bands. The sensitivity verification can reach 100 fg at the DNA level, which is 1000 times of the detection sensitivity of the common PCR method. The LAMP test was carried out on the DNA extracted from the suspected black smut disease strain and its rhizosphere soil samples, and each of the three samples were positive, and the test results of the diseased plants and the tissue separation method were consistent with the detection results of Phytophthora nicotianae. After 2 days of inoculation, Phytophthora nicotianae can be detected in tobacco plants with no obvious symptoms of tripping. The Phytophthora nicotiana LAMP detection system established in this study can quickly and easily detect the Phytophthora nicotianae in the diseased plants and their carrying soil.