8株动物源编码Stx2短尾噬菌体的生物学特性

目的]对8株源自大肠杆菌O157编码Stx2毒素的噬菌体生物学特性进行研究.方法]丝裂霉素C诱导8株大肠杆菌O157菌株释放噬菌体,采用PCR作初步鉴定,分离、纯化噬菌体基因组,随机引物法地高辛(DIG)标记stx2基因片段作为探针,对纯化的噬菌体采用Southernblot进行Stx2噬菌体再次鉴定,透射电子显微镜观察纯化的8株Stx2噬菌体的形态特征,通过限制性内切酶图谱分析,确定噬菌体的核酸类型和基因组大小、以及限制性内切酶酶切片段多态性,并分析噬菌体的蛋白质组成特征.结果]Southern blot证实分离的8株噬菌体为Stx2噬菌体,电镜下观察的各株Stx2噬菌体形态一致,头部均为正六边形,尾部很短,属于短尾噬菌体科,各株噬菌体之间存在相同的蛋白结构模式,基因组为双链DNA,限制性内切酶片段长度表现出一定的多态性,噬菌体的基因组大小从48.0-65.3 kb不等.结论]来源不同菌株的8株编码Stx2噬菌体均为短尾噬菌体,其蛋白结构模式一致,但基因组具有不同组成.

Biologic Characteristics of Eight Podoviridae Stx2-converting phage

Objective] We studied biologic characteristics of Stx2-converting phage induced from Escherichia coli O157 by mitomycin C. Methods] Eight Stx2-converting phages were isolated from E. coli O157 and identified by PCR. The phage particles were purified and phage DNA was extracted. Random priming digoxin (DIG)-labeled stx2-specific gene probe was prepared for Southern blot. The morphology of these phages were studied by electron microscopy. Protein profiles were analyzed by Sodium Dodecyl Sulphate PolyAcrylamide Gel Electrophoresis (SDS-PAGE). Restriction fragment length polymorphism (RFLP) was used to confirm the size, type, and polymorphism of the purified phage genome. Results] These 8 phages were confirmed Stx2-converting phage and DIG-labeled probe was highly specific. All phages had a regular hexagonal head and a short tail, belonging to Podoviridae phage families. The Stx2 phages had genome sizes in the range of 48 to 65.3kb, consisting of dou-ble-stranded DNA. The restriction fragment length polymorphism of these phages showed different groups, although the SDS-PAGE protein profiles of these phages were similar. Conclusion] These 8 Stx2-converting phages with similar morphol-ogy belonged to Podoviridae phage families. The protein profiles were highly identical. We could differentiate these Stx2-converting phages according to their restriction fragment length polymorphism patterns.