| 摘要: |
| 【目的】建立一种定量检测噬菌体宿主菌的简捷方法。【方法】按体积比1∶9将4.4×109 pfu/mL的T4工作液和已知活菌浓度的T4宿主型大肠杆菌工作液混合形成模拟样品,立即在37 ℃ 150 r/min下作用5 min完成侵染,接着采用离心、微滤分离技术清除样品中直径0.22 μm以下的物质(包括游离T4),同时将颗粒物(包括受侵染的大肠杆菌)截留在微滤膜中。待受侵染大肠杆菌裂解后,用SM缓冲液将子代T4从微滤膜中洗滤出来,收集滤液得到转换样品。通过噬菌斑计数法测出转换样品子代T4数量,分析其与模拟样品T4宿主型大肠杆菌活菌数量的定量关系。【结果】模拟样品T4宿主型大肠杆菌活菌数的对数和转换样品子代T4数量的对数呈一元线性关系;参试菌株和试验条件决定着方程的斜率和截距。【结论】按照上述方法转换样品,并根据转换样品子代T4数量和上述一元线性关系,可以快速测定样品中T4宿主型大肠杆菌的活菌含量。 |
| 关键词: 大肠杆菌 T4噬菌体 定量检测 |
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| Initial detection of T4 host E. coli based on sample transformation |
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| Abstract: |
| 【Objective】 A simple and fast method was investigated for quantitative detection of phage host.【Method】 The T4 suspension of 4.4×109 pfu/mL and the T4-host E.coli suspension of known CFU were mixed by 1∶9 (V/V) to make a simulated sample.It was immediately incubated at 37 ℃ with 150 r/min for 5min to complete infection.Then the substances smaller than 0.22μm,including free T4s,were separated by supercentrifugation and millipore filtration.Meanwhile,the particulates,including infected E.colis,were trapped in the syringe filter.After the E.colis lysed,the progeny T4s were eluted out from the filter by SM buffer to get derivative sample.Then progeny T4s were quantitated by plaque counting,and the quantitative relationship between the numbers of the alive T4-host E.colis in the simulated sample and the progeny T4s in the derivative sample was analyzed.【Result】 The logarithm of the progeny T4 number was linear correlated with the logarithm of the T4 host number, shown as an equation of linear regression.The slope rate and the intercept of the equation depended on the tested E.coli strain and the experimental conditions.【Conclusion】 Transforming the sample with the method mentioned above，and relying on the progeny T4 number in the derivative sample and the linear relationship,the alive T4-host E.coli in the sample can be fast enumerated. |
| Key words: E.coli T4 quantitative detection |
