Abstract
In the Czech Republic, three closely related species of the genus Tilletia belong to pathogens that cause significant losses of wheat crops by replacing grains with a mass of teliospores. A quantitative real-time qPCR assay using SYBR Green I has been developed to quantify the amount of T. caries and T. controversa mycelium in apical meristems of different wheat varieties. The real-time PCR reaction was validated by evaluating selected extraction methods, examining the specificity of designed target-specific IGS primers and verifying the optimised amplification reaction on naturally infected wheat plants. The PCR detection limit for the specific identification of fungal DNA was 0.22 ng of mycelium, and the negative correlation between the target DNA quantity and cycle threshold (Ct) was high with a coefficient of determination of R<sup>2</sup> = 0.992. The developed method was used to quantify pathogens mycelium in five wheat varieties in the range from 0.34 ng to 15 &micro;g per one growing tip.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have