Invasive Species Compendium

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Abstract

Development of race specific SCAR marker for the detection of Ralstonia solanacearum (race 2).

Abstract

A PCR based technique has been developed for the detection and identification of Ralstonia solanacearum race 2, the causal organism of bacterial wilt disease. The 114 isolates collected from wilt affected fields of Karnataka and Goa states were studied. Cultural characteristics, pathogenicity test and PCR amplification of 16S rDNA confirmed the identity of R. solanacearum. Based on leaf infiltration test on tobacco leaves, 42 isolates were found to belong to race 2 and 72 to race 3. Polymorphic amplicon of size 600 bp was obtained with OPAB-08 RAPD primer in ten selected race 2 isolates. Identified RAPD marker was converted into SCAR through cloning and sequencing of RAPD amplicon, and the SCAR primers were developed. The SCAR primer pair, RS-R2F and RS-R2R, was used for the identification of race 2 bacteria through specific PCR amplification. Among all the 114 bacterial isolates tested for PCR amplification along with reference strain, only SCAR RS-R2 unambiguously amplified a single amplicon of the original RAPD in 42 race 2 isolates. This SCAR marker, can potentially distinguish race 2 from race 1 and 3 and the technique is rapid, sensitive, easy to perform and feasible for race 2 specific diagnosis of R. solanacearum in wilted plants.