Green Michael R, Sambrook Joseph
Cold Spring Harb Protoc. 2017 Nov 1;2017(11):pdb.prot093435. doi: 10.1101/pdb.prot093435.
Stocks of bacteriophage M13 are usually grown in liquid culture. The infected bacteria do not lyse but, instead, grow at a slower than normal rate to form a dilute suspension. The inoculum of bacteriophage is almost always a freshly picked plaque or a suspension of bacteriophage particles obtained from a single plaque, as described here. Infected cells contain up to 200 copies of double-stranded, replicative-form DNA and extrude several hundred bacteriophage particles per generation. Thus, a 1-mL culture of infected cells can produce enough double-stranded viral DNA (1-2 mg) for restriction mapping and recovery of cloned DNA inserts and sufficient single-stranded DNA (∼5-10 mg) for site-directed mutagenesis, DNA sequencing, or synthesis of radiolabeled probes. The titer of bacteriophages in the supernatant from infected cells is so high (∼10 pfu/mL) that a small aliquot serves as a permanent stock of the starting plaque.
噬菌体M13的储备液通常在液体培养物中培养。被感染的细菌不会裂解，而是以比正常速度慢的速度生长，形成稀释的悬浮液。噬菌体的接种物几乎总是如本文所述的新鲜挑取的噬菌斑或从单个噬菌斑获得的噬菌体颗粒悬浮液。被感染的细胞含有多达200份双链复制型DNA，并且每代挤出数百个噬菌体颗粒。因此，1 mL被感染细胞的培养物可以产生足够的双链病毒DNA（1-2 mg）用于限制性图谱分析和克隆DNA插入片段的回收，以及足够的单链DNA（约5-10 mg）用于定点诱变、DNA测序或放射性标记探针的合成。感染细胞上清液中噬菌体的滴度非常高（约10 pfu/mL），以至于一小份就可作为起始噬菌斑的永久储备液。
