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目的从纳帕海湿地土样中分离和培养低温菌及其噬菌体,并对其特性进行研究。方法分离纯化低温宿主菌,采用“双层平板法”分离获得噬菌体,通过16S rRNA基因分析对宿主菌进行初步鉴定,对噬菌体进行电镜形态观察,并进行噬菌体基因组酶切片段长度多态性分析及噬菌体生理特性研究。结果从纳帕海土样中分离获得一株裂解性低温噬菌体,命名为SV-12,其宿主菌S-12鉴定为恶臭假单胞菌(Pseudomonas putida)。噬菌体SV-12有囊膜,头部为典型的正六边形,立体对称结构,直径约68.7 nm;尾管长约122.5 nm,直径约25 nm;在4℃时具有侵染活性,4～25℃均能产生边缘清晰、透明的噬菌斑。最适感染温度约为10℃,pH耐受范围较广,在pH=10时具有最多出斑数。结论 SV-12属于肌尾噬菌体科,为裂解性噬菌体,对氯仿极度敏感。基因组为dsDNA,大小约为74 kb。
OBJECTIVE To isolate and cultivate the thermophilic bacteria and their phage from the Napahai wetland soil samples and study their characteristics. Methods The phage were isolated and purified from the host cells under low temperature conditions. The phage were obtained by the “double-layer plate method”. The 16S rRNA gene analysis was used to identify the host bacteria. The phage were observed by electron microscopy and the phage genome fragment length polymorphism Analysis and Research on Physiological Characteristics of Phage. Results A lytic bacteriophage was obtained from Napahai soil samples and named SV-12. Its host strain S-12 was identified as Pseudomonas putida. Phage SV-12 has a capsule membrane, the head is a typical hexagonal, three-dimensional symmetrical structure, a diameter of about 68.7 nm; tail tube length of about 122.5 nm, a diameter of about 25 nm; 4 at infection activity, 4-25 ℃ can produce clear edge, transparent plaque. Optimal infection temperature is about 10 ℃, pH tolerance a wider range, at pH = 10 with the largest number of spots. Conclusion SV-12 belongs to myofibroblastomycetes and is a lytic bacteriophage, which is extremely sensitive to chloroform. The genome is dsDNA, about 74 kb in size.