Development and Optimization of Molecular Technique for Diagnosis of Citrus Canker in Citrus Cultivars
Development and Optimization of Molecular Technique for Diagnosis of Citrus Canker in Citrus Cultivars
JAVERIA NEHAL1, ARIF MUHAMMAD KHAN1, AZHAR ABBAS KHAN*2, MUHAMMAD MUBIN3, HAFIZ MUHAMMAD TAHIR4 & JAVED IQBAL5
ABSTRACT
The conventional method for the detection of Xanthomonas axonopodis has been based on biochemical tests. A rapid and sensitive method for identification and detection of Xanthomonas axonopodis is required for management of citrus canker. PCR-based diagnostic test is appropriate for monitoring pathogen in a very short time compared to laborious, non-specific and expensive protocols. ELISA, Nested PCR has been used for many years in different countries. Citrus orchards in Sargodha region were surveyed and leaf samples showing typical symptoms of citrus canker were collected. Infected section of leaf was taken for isolation of bacteria. Lesions were cut into parts and streak bacteria by the help of inoculation loop grown in nutrient medium. Pure bacterial culture of Xanthomonas axonopodis were used for detection by standard PCR. Xanthomonas axonopodis was diagnosed by amplification of 16S rDNA. A fragment of ~1.4 kb was amplified and cloned for sequencing Out of three markers used (K1F-CIT/ K1R-CIT, K2F-CIT/ K2R-CIT, K3F-CUT/ K3R-CIY) K3F-CUT/ K3R-CIY gave best results repeatedly. So this primer pair can be used for identification/diagnosis of Xanthomonas axonopodis. Bacterial culture used as template in PCR and colony PCR gave better results as compared with extracted DNA from infected leaf.
Key Words: Xanthomonas axonopodis citri, Citrus canker, Molecular diagnostics
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December 2017
Vol.63, Iss. 2, Pages 125-234
