Preserved Microarrays for Simultaneous Detection and Identification of Six Fungal Potato Pathogens with the Use of Real-Time PCR in Matrix Format

Biosensors

Vitaly Dzhavakhiya¹  Natalya Girsova¹  Mikhail Pridannikov¹  Natalia Statsyuk¹  Maria Kuznetsova¹  Alexander Golikov²  Maksim Nikitin²  Ksenia Deych²  Inessa Grevtseva² 

[1] All-Russian Research Institute of Phytopathology, Institute Str., 5, Bolshie Vyazemy 143050, Russia;GenBit LLC, Nauchny pr., 20, Bld. 4, Moscow 117246, Russia;

关键词: plant pathogen diagnostics;    qPCR microarrays;    potato;    Spongospora subterranea;    Rhizoctonia solani;    Alternaria solani;    Alternaria alternata;    Colletotrichum coccodes;    Fusarium spp.;   

DOI  :  10.3390/bios8040129

来源: DOAJ

【 摘 要 】

Fungal diseases of plants are of great economic importance causing 70⁻80% of crop losses associated with microbial plant pathogens. Advanced on-site disease diagnostics is very important to maximize crop productivity. In this study, diagnostic systems have been developed for simultaneous detection and identification of six fungal pathogens using 48-well microarrays (micromatrices) for qPCR. All oligonucleotide sets were tested for their specificity using 59 strains of target and non-target species. Detection limit of the developed test systems varied from 0.6 to 43.5 pg of DNA depending on target species with reproducibility within 0.3−0.7% (standard deviation). Diagnostic efficiency of test systems with stabilized and freeze-dried PCR master-mixes did not significantly differ from that of freshly prepared microarrays, though detection limit increased. Validation of test systems on 30 field samples of potato plants showed perfect correspondence with the results of morphological identification of pathogens. Due to the simplicity of the analysis and the automated data interpretation, the developed microarrays have good potential for on-site use by technician-level personnel, as well as for high-throughput monitoring of fungal potato pathogens.

【 授权许可】

Unknown