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PCR test for the presence of Ralstonia solanacearum

UH Hilo Home > Academics > College of Agriculture, Forestry and Natural Resource Management > Research > Ralstonia solanacearum

gel electropherogram showing PCR results from ginger samples The samples on this gel are from left, A, B, C, and D, followed by a lousy ladder. Sample B is negative, and the rest are positive. See pictures of the rhizomes.

This test is based on the selective amplification of the 16S ribosomal rRNA gene from R. solanacearum (Seal and Jackson, 1991). Basically if a fragment gets amplified and shows up as a band on a gel, R. solanacearum was present in the sample.

It's hard to tell from the above gel, but the amplified fragment is approximately 300 bp. long, and comes from a PCR with the primers OLI1 and Y2.

Below you see a diagram of the 16s rRNA gene, which is very highly conserved among bacteria. The primers Y1 and Y2 are considered "universal" in that they amplify this gene from most eubacteria.

cartoon showing the region of DNA amplified by the Y1, Y2 and OLI1 primers
Click for larger view.

The OLI1 primer targets a region of heterogeneity which differentiates Rs from other bacteria. Thus, when OLI1 and Y2 are used in PCR, amplification only occurs when Rs is present.