Fig. 1. Agarose gel electrophoresis (left) and visual detections (right) after loop-mediated isothermal amplification (LAMP) reaction of DNA samples with the LAMP primer set (WBPH-65). In the agarose gel electrophoresis, various size of bands, such as ladder, were observed in the PC and 100 ng of WBPH DNA only, but no band in the NC and single bands in 100 ng of BPH DNA and 100 ng of SBPH DNA were observed. This indicates that the WBPH-65 LAMP primer set designed in this study can amplify the target region in the genomic DNA of WBPH specifically, but not in the genomic DNA of BPH and SBPH. Visual detections showed that the WBPH-65 LAMP primer set was also able to specifically amplify WBPH genomic DNA with a sensitivity of 0.01 ng in minimum, but not in the two rice planthoppers, BPH and SBPH. For LAMP reaction, all samples were incubated at 65℃ for 60 min and enzyme was inactivated at 80℃ for 5 min. †M, 100 bp ladder marker; PC, positive control (LAMP primer set and DNA, both were provided by manufacturer); NC, negative control (WBPH-65 LAMP primer set without DNA); WBPH, white-backed planthopper Sogatella furcifera; BPH, brown planthopper Nilaparvata lugens; SBPH, small brown planthopper Laodelphax striatellus.
Fig. 2. Visual detections after loop-mediated isothermal amplification (LAMP) reaction with the LAMP primer set (WBPH-65) according to the incubation time (20, 30, 40, and 60 min) and the amount of genomic DNA (0, 0.01, 0.1, 1, 10, and 100 ng) of white-backed planthopper (WBPH) Sogatella furcifera. After the LAMP reaction, enzyme within the tube was inactivated at 80℃ for 5 min.
Fig. 3. Loop-mediated isothermal amplification (LAMP) reaction results on five individuals of three rice planthoppers, white-backed planthopper (WBPH; Sogatella furcifera), brown planthopper (BPH; Nilaparvata lugens), and small brown planthopper (SBPH; Laodelphax striatellus) with two LAMP primer sets of WBPH-65, [F3 + B3 + FIP + BIP + LF + LB] and [F3 + B3 + FIP + BIP], respectively. The incubation time was 60 min at 65℃ and the amount of genomic DNA of each planthopper was > 100 ng. After the LAMP reaction, a Bst polymerase was inactivated at 80℃ for 5 min.
Table 1. Loop-mediated isothermal amplification (LAMP) primer sets designed in this study to discriminate Sogatella furcifera from other planthopper species