The species Puccinia triticina (Pt) and Puccinia striiformis f. sp. tritici (Pst) are devastating cereal pathogens that cause leaf and stripe rust diseases. We developed PCR assays for the species-specific detection of Pt and Pst, 2 biological agents that cause wheat rust disease. For each pathogen, we validated 3 primer sets that target the second largest subunits of the RNA polymerase II (rpb2) and β-tubulin 1 (tub1) genes. The specificities of the primers were verified using naturally infected plant materials with visual symptoms of disease. All primer sets amplified a single DNA fragment of the expected length. The primer sets LidPr15/16, LidPr1/2, and LidPs13/14 were able to detect small amounts of pure fungal DNA with sensitivities of 0.1, 1, and 10 pg/μL, respectively. A sufficient detection limit (1 pg/μL to 5 ng/μL) was observed for all assays when the sensitivity test was performed with host plant DNA. The study also evaluated the simultaneous detection of both rust pathogens, and the multiplex PCR assay generated amplicons of 240 and 144 bp in length for Pts (LidPs9/10) and Pt (LidPr1/2), respectively.

中文翻译：

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新型PCR检测小麦锈病的生物因子的检测新PCR：小麦锈菌和小麦锈菌。sp。小麦。

小麦Puccinia triticina（Pt）和Puccinia striiformis f.。sp。Tritici（Pst）是破坏性的谷物病原体，可引起叶和条锈病。我们开发了用于检测Pt和Pst（两种引起小麦锈病的生物制剂）的物种特异性检测的PCR分析方法。对于每种病原体，我们验证了针对RNA聚合酶II（rpb2）和β-微管蛋白1（tub1）基因第二大亚基的3个引物对。使用具有疾病视觉症状的天然感染植物材料验证了引物的特异性。所有引物组均扩增了预期长度的单个DNA片段。引物对LidPr15 / 16，LidPr1 / 2和LidPs13 / 14能够分别检测少量纯真菌DNA，其灵敏度分别为0.1、1和10 pg /μL。使用寄主植物DNA进行敏感性测试时，所有测定均观察到足够的检测限（1 pg /μL至5 ng /μL）。该研究还评估了两种锈病病原体的同时检测，并且多重PCR分析分别生成了长度分别为Pts（LidPs9 / 10）和Pt（LidPr1 / 2）的240和144 bp的扩增子。