A positive reaction was indicated by a colour change from violet to sky blue (Figs 2c, 3b and 4c). The LAMP reaction with HNB could also be performed in a 96-well microplate (Goto et al., 2009) and would be helpful for high-throughput DNA detection. Meanwhile, the positive reactions by self-trial were seen as a ladder-like pattern on 2% agarose gel electrophoresis analysis, verifying the results of the visual detection with HNB (Figs 2b and 4b). The detection limit of P. sojae using the three methods was 10 pg μL−1 (Fig. 4).
This is in accordance with two reports on LAMP methods used to detect Phytophthora spp. (Tomlinson et al., 2007, Z-VAD-FMK molecular weight 2010). Moreover, it has been reported that the LAMP reaction might be facilitated by the addition of loop forward and backward primers (Nagamine et al., 2002). In the present study, we could not identify a suitable loop forward primer, so we only used the loop backward primer to accelerate
the reaction (Table 1). This improved the reaction time by approximately 10-fold (data not shown). In the field trial, we collected 130 diseased soybean tissues and residues. All samples were inspected by LAMP, PCR, and a leaf disk-baiting method for comparison (Table 2). Compared with the other methods, the newly developed A3aPro-LAMP significantly improved the detection efficiency. Thus, the A3aPro-LAMP assay developed in this study can be used for the rapid diagnosis of P. sojae KU-60019 order in plants and in production fields. This, in turn, many could make it possible to control the dispersion of P. sojae and increase Phytophthora-free soybean production. This research was supported by the National Department Public Benefit Research Foundation (No. 200903004), the National ‘863’ Program (2012AA101501), the ‘948’ project (2010-C17) and Chinese National Science Foundation Committee project (3-20). We thank Michael D. Coffey from University of California Riverside for providing us with an isolate of Phytophthora vignae. “
“The EngA protein is a conserved and essential
bacterial GTPase of largely enigmatic function. While most investigations of EngA have suggested a role in ribosome assembly, the protein has also been implicated in diverse elements of physiology including chromosome segregation, cell division, and cell cycle control. Here, we have probed additional phenotypes related to ribosome biogenesis on depletion of EngA in Escherichia coli to better understand its role in the cell. Depletion of EngA resulted in cold-sensitive growth and stimulation of a ribosomal rRNA promoter, both phenotypes associated with the disruption of ribosome biogenesis in bacteria. Among antibiotics that inhibit translation, depletion of EngA resulted in sensitization to the aminoglycoside class of antibiotics. EngA bound the alarmone ppGpp with equally high affinity as it bound GDP. These data offer additional support for a role in ribosome biogenesis for EngA, possibly in maturation of the A-site of the 50S subunit.