Molecular detection of Fusarium oxysporum f. sp. vasinfectum in cotton roots by PCR and real-time PCR assay Molekularer Nachweis von Fusarium oxysporum f. sp. vasinfectum in Baumwolle mittels PCR und Real-Time-PCR

A PCR assay based on a pair of oligonucleotide primers targeting the 16S and 23S rRNA genes was used to detect Fusarium oxy- sporum f. sp. vasinfectum (Fov), a fungus causing Fusarium wilt of cotton, in infected cotton seedlings. By using the primer pair Fov1-Eg-f and Fov1-Eg-r, a 438-bp DNA fragment was consistent- ly amplified from 41 Egyptian isolates of Fov (race 3), while no amplification was obtained from template genomic DNAs ob- tained from other fungal pathogens, namely F. oxysporum, F. solani, F. verticillioides, F. sambucinum, Rhizoctonia solani, and Macrophomina phaseolina. PCR product of the expected size was amplified from as little as 100 fg of purified Fov genomic DNA. The sensitivity of real-time PCR allowed detection of 200 fg of target genomic DNA. The melting temperature (Tm) values of Fov isolates were in the range 86.83 to 87.02. Fov DNA was de- tected in infected cotton roots, while no amplification was ob- tained from other plant structures, such as stem and leaf. The re- al-time PCR assay successfully detected Fov and correlated well with assessments based on conventional PCR techniques. The application of this technique will be useful for identification of Fov in infected cotton roots and detection of Fov in infected plants that lack visible morphological structures or symptoms.