Filamentous bacteriophages are widely used in phage display technology. The most common quantification method is lysis plaque formation test (PFT). This technique has several disadvantages, and only quantifies infective phages and is not effective when phagemids are used. We developed a qPCR method directed against the M13 replication origin, which detects between 3.3 × 10³ and 3.3 × 10⁸ viral genome copies with a linearity of R² = 0.9998. Using this method we were able to observe a difference of approximately ten more phages than with the PFT. This difference was not due to the presence of a free genome, which suggests the presence of non-infective particles. Using a DNaseI treatment, we observed the presence of 30% to 40% of unpackaged genome in recombinant phage modified in PIII or PVIII. The qPCR method with a DNase I treatment is an efficient method to quantify the total amount of filamentous phages.

中文翻译：

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针对病毒复制起点的qPCR设计用于量化丝状噬菌体和噬菌粒的总量。

丝状噬菌体广泛用于噬菌体展示技术。最常见的定量方法是溶菌斑形成试验（PFT）。该技术具有几个缺点，并且仅定量感染噬菌体，并且在使用噬菌粒时无效。我们开发了针对M13复制起点的qPCR方法，该方法可检测线性范围为R ^2的3.3×10 ³至3.3×10 ⁸病毒基因组拷贝 = 0.9998。使用这种方法，我们可以观察到比PFT多大约十个噬菌体的差异。这种差异不是由于存在游离基因组，这表明存在非感染性颗粒。使用DNaseI处理，我们观察到在PIII或PVIII中修饰的重组噬菌体中存在30％至40％的未包装基因组。用DNase I处理的qPCR方法是定量丝状噬菌体总量的有效方法。