Detection of Tobacco Bacterial Wilt Caused by Ralstonia solanacearum by Combining Polymerase Chain Reaction with an α-Hemolysin Nanopore

Abstract

Tobacco bacterial wilt is a serious disease caused by the soil-borne bacterium Ralstonia solanacearum (R. solanacearum). Herein, a rapid and purification-free α-hemolysin (α-HL) nanopore-sensing strategy based on polymerase chain reaction (PCR) and lambda exonuclease digestion was established to detect R. solanacearum. A 198-nucleotide-long single-stranded DNA was obtained via asymmetric PCR or the lambda exonuclease-mediated digestion of the PCR product. The DNA fragment produced unique long-lived, current-blocking signals when it passed through the α-HL nanopore. This sensing approach can allow for the determination of R. solanacearum in tobacco samples and can be conveniently extended to other DNA monitoring because of the extremely wide range of PCR applications.