Chasteen L, Ayriss J, Pavlik P, Bradbury A R M
B Division, Los Alamos National Laboratory MS M888, Los Alamos, NM 87545, USA.
Nucleic Acids Res. 2006;34(21):e145. doi: 10.1093/nar/gkl772. Epub 2006 Nov 6.
Phage display technology involves the display of proteins or peptides, as coat protein fusions, on the surface of a phage or phagemid particles. Using standard technology, helper phage are essential for the replication and assembly of phagemid particles, during library production and biopanning. We have eliminated the need to add helper phage by using 'bacterial packaging cell lines' that provide the same functions. These cell lines contain M13-based helper plasmids that express phage packaging proteins which assemble phagemid particles as efficiently as helper phage, but without helper phage contamination. This results in genetically pure phagemid particle preparations. Furthermore, by using constructs differing in the form of gene 3 that they contain, we have shown that the display, from a single library, can be modulated between monovalent (phagemid-like) and multivalent display (phage-like) without any further engineering. These packaging cells eliminate the use of helper phage from phagemid-based selection protocols; reducing the amount of technical preparation, facilitating automation, optimizing selections by matching display levels to diversity, and effectively using the packaged phagemid particles as means to transfer genetic information at an efficiency approaching 100%.
噬菌体展示技术是指将蛋白质或肽作为外壳蛋白融合体展示在噬菌体或噬菌粒颗粒表面。使用标准技术时，在文库构建和生物淘选过程中，辅助噬菌体对于噬菌粒颗粒的复制和组装至关重要。我们通过使用具有相同功能的“细菌包装细胞系”消除了添加辅助噬菌体的需求。这些细胞系含有基于M13的辅助质粒，可表达噬菌体包装蛋白，这些蛋白组装噬菌粒颗粒的效率与辅助噬菌体相同，但不会受到辅助噬菌体污染。这使得制备出的噬菌粒颗粒基因纯净。此外，通过使用含有不同形式基因3的构建体，我们表明，从单个文库中展示可以在单价（类似噬菌粒）和多价展示（类似噬菌体）之间进行调节，而无需任何进一步的工程改造。这些包装细胞在基于噬菌粒的筛选方案中无需使用辅助噬菌体；减少了技术准备工作量，便于自动化操作，通过使展示水平与多样性相匹配来优化筛选，并有效地将包装好的噬菌粒颗粒用作传递遗传信息的手段，效率接近100%。
