Specific primers of Paraphoma radicina which causes alfalfa Paraphoma root rot

Autor: Shu Zhong Dang, Jin Ling Hu, Yan Zhong Li, Shi Cao
Rok vydání: 2021
Předmět:
Zdroj: European Journal of Plant Pathology. 162:489-499
ISSN: 1573-8469
0929-1873
Popis: Paraphoma radicina is the causal pathogen of alfalfa root rot. Identification of P. radicina using the traditional morphological methods is difficult and time-consuming. Therefore, there is a need to develop rapid molecular techniques that can detect and identify P. radicina. This study employed conventional and real-time polymerase chain reaction (PCR) assays using a primer pair designed from the pathogen’s internal transcribed spacer (ITS) region to detect the P. radicina levels in alfalfa plants. Accordingly, a 363 bp fragment of P. radicina DNA was amplified using the primers Par-F and Par-R. Conventional PCR could detect as little as 1.2 pg/μL, but real-time PCR was more sensitive and could amplify as low as 12 fg/μL. The two PCR methods detected P. radicina from 15 alfalfa field plants, among which nine had typical P. radicina symptoms, while the other six did not display the typical symptoms. These results show that the PCR assays were more sensitive in detecting P. radicina than the traditional isolation methods. Moreover, relative qPCR, employing the comparative threshold cycle (Ct) method, was reliable for quantifying P. radicina DNA from the infected alfalfa root tissues. The P. radicina DNA quantities detected by real-time PCR assay had high correlations with the disease severity index (DSI) rating of alfalfa cultivars. This study is the first report of P. radicina identification by conventional and real-time PCR assays. The assays might be useful in detecting P. radicina in alfalfa field plants and determining the levels of P. radicina DNA in different alfalfa cultivars for pathogen resistance evaluation.
Databáze: OpenAIRE
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