Generation of affinity-tagged fluoromycobacteriophages by mixed assembly of phage capsids.
Generation of affinity-tagged fluoromycobacteriophages by mixed assembly of phage capsids.
复制标题
通过噬菌体衣壳的混合组装生成亲和标记的荧光分枝杆菌噬菌体。
DOI：
10.1128/aem.01016-13
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发表时间：
2013
影响因子：
4.4
通讯作者：
Hatfull,GrahamF
中科院分区：
文献类型：
--
作者：
Piuri,Mariana;Rondón,Liliana;Urdániz,Estefanía;Hatfull,GrahamF
向噬菌体颗粒中添加亲和标签有助于多种应用，包括疫苗构建和细菌感染的诊断。向噬菌体衣壳添加标签是期望的，因为尾部的修饰可导致吸附差和感染性丧失。虽然标签可以容易地作为融合物包括到头部装饰蛋白，但许多病毒颗粒不具有作为病毒体组分的装饰蛋白。在分枝杆菌噬菌体TM 4衣壳亚基gp 9中加入一个小的（10个氨基酸）Strep-标签II（STAG II）作为遗传上同源的重组噬菌体是不被容忍的，但可以通过在表达衣壳蛋白-STAG融合的宿主上生长野生型噬菌体而被掺入头部。具有由野生型和STAG标记的亚基混合物组成的衣壳的颗粒可以生长到高滴度，显示出良好的感染性，并且可以用于分离噬菌体-细菌复合物。STAG标记的荧光菌噬菌体的制备使得能够捕获细菌复合物并通过荧光鉴定感染的细菌。
Addition of affinity tags to bacteriophage particles facilitates a variety of applications, including vaccine construction and diagnosis of bacterial infections. Addition of tags to phage capsids is desirable, as modification of the tails can lead to poor adsorption and loss of infectivity. Although tags can readily be included as fusions to head decoration proteins, many phages do not have decoration proteins as virion components. The addition of a small (10-amino-acid) Strep-tag II (STAG II) to the mycobacteriophage TM4 capsid subunit, gp9, was not tolerated as a genetically homogenous recombinant phage but could be incorporated into the head by growth of wild-type phage on a host expressing the capsid-STAG fusion. Particles with capsids composed of wild-type and STAG-tagged subunit mixtures could be grown to high titers, showed good infectivities, and could be used to isolate phage-bacterium complexes. Preparation of a STAG-labeled fluoromycobacteriophage enabled capture of bacterial complexes and identification of infected bacteria by fluorescence.