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Tuesday, 20 Aug
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Development of Fusarium oxysporum species complex (FOC) specific primers using Topoisomerase-II gene

 

Abstract

Fusarium is widely regarded as one of the most destructive and species-rich groups of phytopathogens. Most of the important crops affected by Fusarium leads to great loss to global agricultural economy. Members of the Fusarium oxysporum complex (FOC) are common soil-borne fungi which are known to cause wilt, crown rot and root rot in a number of crops. Therefore, accurate identification of the FOC pathogens is required for disease management in crops.In this study,75 Fusarium isolates were recovered from 20 wilted plant and soil samples were taken from NBAIM fields and identified using conventional morphological techniques. Twelve representative strains ofFusarium spp. were identified using 28SrDNA and topoisomerase-II (topo-II) gene sequencing. Sequence analysis of both the genes were made by examining conserved sequences within species and variable sequence in different species in which no species specific sequences were found in 28S rDNA but, were present in topo-II gene sequences. FOC specific forward primer (FO­-F) and reverse primer (FO-R) were designed by selecting the specific regions of topo-II gene sequence and tested with five F. oxysporum isolates along with some non-F. oxysporum isolates. The results revealed a single band size of 495 bp in all tested F. oxysporum isolates which was absent in other isolates. The specificity of the primer was validated by taking 23 Fusarium isolates from wilted plants and soil samples. The results showed positive amplification in Fusarium oxysporum isolates whereas no amplification observed in related species.  The developed primers were able to identify and discriminate FOC from related species and can be used in ecological studies, disease diagnostics and in screening of infected plants.

Key Words

Fusarium oxysporum, PCR, topoisomerase-II gene, 28SrDNA

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