枯草芽胞杆菌PTS-394的GFP标记及其定殖能力

为探明枯草芽胞杆菌PTS-394在番茄根围的定殖能力，采用电转化法获得绿色荧光蛋白（green fluorescent protein，GFP）标记菌株PTS-GFP，构建其生长曲线，采用对峙生长法评价其室内抑菌活性，并应用抗生素平板回收结合激光扫描共聚焦显微镜观察标记菌株在番茄根围的定殖数量。结果显示：与原始菌株PTS-394相比，标记菌株PTS-GFP的生长、对青枯病菌和4种病原真菌的室内抑菌能力无明显差异。标记菌株PTS-GFP和青枯病菌菌液单独或混合处理番茄苗，灌根当天标记菌株初始菌量接近10⁸ CFU/g，处理3 d后种群数量迅速下降，约10⁶ CFU/g，随后缓慢下降，处理10 d后种群数量约10⁴ CFU/g，处理30 d后，标记菌株仍然能被检测到，约20 CFU/g。表明枯草芽胞杆菌PTS-394在番茄根际土壤中具有一定的定殖能力。

Bacillus subtilis PTS-394 labeled by green fluorescent protein and its colonization

To investigate the colonization of Bacillus subtilis PTS-394 in the rhizosphere of tomato, the GFP-labeled strain PTS-GFP was obtained by electroporating the shuttle vector pGFP22, harboring the gfp gene, into the original strain PTS-394. Dynamic growth analyses and antimicrobial activities in vitro of PTS-GFP were tested in this study. PTS-GFP colonization in the rhizosphere of tomato was conducted by combining antibiotics plate recovery and laser scanning confocal microscope. The results showed that there was no obvious difference in the cell growth and inhibition against the tested pathogens between original strain PTS-394 and transformant PTS-GFP. The population of PTS-GFP on the roots of tomato reached 10⁸ CFU/g after inoculating PTS-GFP alone or combined with Ralstonia solanacearum, and then decreased sharply 3 days later, at 10⁶ CFU/g, and remained at 10⁴ CFU/g 10 days later. PTS-GFP in the rhizosphere of tomato could survive for over 30 days, and eventually remained at 20 CFU/g. The results indicated that B.subtilis PTS-394 displayed good colonization ability in the rhizosphere of tomato.