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Abstract
The present study reports the development and evaluation of a hemi-nested polymerase chain reaction (hnPCR) assay for the efficient detection of ‘Candidatus Liberibacter’ infecting citrus plants. A set of new primers was designed by alignment of nucleotide sequences of the β-operon (rplKAJL-rpoBC) ribosomal protein genes from all the known ‘Candi-datus Liberibacter asiaticus’ isolates reported in Genbank. Hemi-nested PCR reaction components and thermal cycling parameters were optimized and reaction conditions were standardized. Sequencing of the PCR products from hemi nested-PCR reactions confirmed the specificity of new primer pairs for ‘Candidatus Liberibacter asiaticus’. The reliability and sensitivity of hnPCR was evaluated by comparing it to real-time PCR.
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