A rapid colorimetric LAMP assay for detection of Rhizoctonia solani AG-1 IA causing sheath blight of rice.
A rapid colorimetric LAMP assay for detection of Rhizoctonia solani AG-1 IA causing sheath blight of rice.
复制标题
一种快速的比色灯测定法，用于检测溶质溶胶的根源，导致米鞘浸泡。
DOI：
10.1038/s41598-020-79117-0
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发表时间：
2020-12-16
影响因子：
4.6
通讯作者：
Saxena AK
中科院分区：
文献类型：
--
作者：
Choudhary P;Rai P;Yadav J;Verma S;Chakdar H;Goswami SK;Srivastava AK;Kashyap PL;Saxena AK
立枯丝核菌是最具破坏性的病原菌之一。R. Solani AG-11 A引起稻、玉米和其它禾本科植物的纹枯病。准确鉴定是有效管理这种病原体的关键。本研究针对植物病原真菌的重要毒力因子多聚半乳糖醛酸酶（PG）基因设计了4对引物，即RSPG 1、RSPG 2、RSPG 4和RSPG 5。4对引物均能扩增出300 bp（RSPG 1F/R）、375 bp（RSPG 2F/R）、500 bp（RSPG 4F/R）和336 bp（RSPG 5 F/R）的扩增子。使用每个引物组的q-PCR检测可以检测高达10 pg的DNA。同时设计了6对引物（RS_pg_F3_1/RS_pg_B3_1、RS_pg_FIP_1.1/RS-pg_BIP_1.1和RS_pg_LF_1/RS_pg_LB_1）。此外，当与来自感染样品的快速高通量模板制备方法（rHTTP）结合时，开发的比色LAMP测定在样品收集的45分钟内产生病原体的视觉确认。LAMP法的灵敏度低至1.65 fg/μl模板DNA，可有效检测R. solani AG-1 IA从患病植物组织和土壤样品中分离。LAMP法对R. solani，因为它与R. solani和密切相关的真菌和细菌外类群。这项研究将有助于设计一种有效的床旁诊断方法，用于早期监测R。solani，从而制定针对病原体的及时预防措施。
Rhizoctonia solani is one of the most devastating pathogens. R. solani AG-1 IA causes sheath blight in rice, maize, and other Gramineous plants. Accurate identification is essential for the effective management of this pathogen. In the present study, a set of four primers were designed viz. RSPG1, RSPG2, RSPG4, and RSPG5 for polygalacturonase (PG) gene, an important virulence factor in phytopathogenic fungi. All four primer sets showed specific amplification of 300 bp (RSPG1F/R), 375 bp (RSPG2F/R), 500 bp (RSPG4F/R) and 336 bp (RSPG5F/R) amplicons. q-PCR detection using each primer sets could detect up to 10 pg of DNA. We also designed six primers (RS_pg_F3_1/RS_pg_B3_1, RS_pg_FIP_1.1/RS-pg_BIP_1.1, and RS_pg_LF_1/RS_pg_LB_1) for PG gene. Further, a colorimetric LAMP assay developed yielded visual confirmation of the pathogen within 45 min of sample collection when coupled with rapid high throughput template preparation method (rHTTP) from infected samples. The sensitivity of the LAMP assay was as low as 1.65 fg/µl of template DNA and could effectively detect R. solani AG-1 IA from diseased plant tissues and soil samples. The LAMP assay was highly specific for R. solani as it did not show any amplification with other AG groups of R. solani and closely related fungal and bacterial outgroups. This study will help in designing an effective point of care diagnostic method for early monitoring of R. solani and thereby planning timely preventive measures against the pathogen.
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影响因子：
4.6
作者：
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通讯作者：
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