Upscaled CTAB-Based DNA Extraction and Real-Time PCR Assays for Fusarium culmorum and F. graminearum DNA in Plant Material with Reduced Sampling Error

International Journal of Molecular Sciences

Christoph Brandfass¹ 

[1] id="af1-ijms-09-02306">University of Göttingen, Department of Crop Sciences, Molecular Phytopathology and Mycotoxin Research Division, Grisebachstrasse 6, 37077 Göttingen, Germa

关键词: Fusarium graminearum Schwabe;    Fusarium culmorum W.G. Smith;    real-time PCR;    DNA extraction;    sampling error;    Fusarium head blight;   

DOI  :  10.3390/ijms9112306

来源: mdpi

【 摘 要 】

Fusarium graminearum Schwabe (Gibberella zeae Schwein. Petch.) and F. culmorum W.G. Smith are major mycotoxin producers in small-grain cereals afflicted with Fusarium head blight (FHB). Real-time PCR (qPCR) is the method of choice for species-specific, quantitative estimation of fungal biomass in plant tissue. We demonstrated that increasing the amount of plant material used for DNA extraction to 0.5–1.0 g considerably reduced sampling error and improved the reproducibility of DNA yield. The costs of DNA extraction at different scales and with different methods (commercial kits versus cetyltrimethylammonium bromide-based protocol) and qPCR systems (doubly labeled hybridization probes versus SYBR Green) were compared. A cost-effective protocol for the quantification of F. graminearum and F. culmorum DNA in wheat grain and maize stalk debris based on DNA extraction from 0.5–1.0 g material and real-time PCR with SYBR Green fluorescence detection was developed.

【 授权许可】

CC BY   
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