文献类型： 外文期刊

作者： Song, Pei-ling ¹ ; Jedryczka, Malgorzata ² ; Irzykowski, Witold ² ; Yan, Meng-jiao ³ ; Huangfu, Hai-yan ³ ; Hao, Li ¹ ;

作者机构： 1.Inner Mongolia Univ, Sch Life Sci, Hohhot 010020, Peoples R China

2.Polish Acad Sci, Inst Plant Genet, Ul Strzeszynska 34, PL-60479 Poznan, Poland

3.Inner Mongolia Acad Agr & Anim Husb Sci, Inst Plant Protect, Hohhot 010031, Peoples R China

关键词： Brassica napus;contaminated seed;detection;Leptosphaeria maculans;oilseed rape

期刊名称：JOURNAL OF PHYTOPATHOLOGY （ 影响因子：1.789； 五年影响因子：1.574 ）

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收录情况： SCI

摘要： An efficient DNA extraction protocol and polymerase chain reaction (PCR) assay for detecting Leptosphaeria maculans from infected seed lots of oilseed rape were developed. L.maculans, the causal agent of blackleg, a damaging disease in oilseeds rape/canola worldwide, was listed as a quarantine disease by China in 2009. China imports several millions of tons of oilseeds every year. So there is a high risk that this pathogen will be introduced to China via contaminated seeds. Seed contamination is one of the most significant factors in the global spread of phytopathogens. Detection of L.maculans in infected seed lots by PCR assay is difficult due to the low level of pathogen mycelium/spores on seeds and PCR inhibitors associated with the seeds of oilseed rape. In our study, these two major obstacles were overcome by the development of a two-step extraction protocol combined with a nested PCR. This extraction protocol (kit extraction after CTAB method) can efficiently extract high-quality DNA for PCR. Amplification results showed that the detection threshold for conventional PCR and nested PCR was, respectively, 1ng and 10fg of DNA per l in mycelia samples. On contaminated seed lots of oilseed rape, the detection threshold of conventional and nested PCR was 709fg/l and 709ag/l of DNA, respectively. The DNA extraction protocol and PCR assay developed in this study can be used for rapid and reliable detection of L.maculans from infected seeds of oilseed rape.