Syvanen M
Proc Natl Acad Sci U S A. 1974 Jun;71(6):2496-9. doi: 10.1073/pnas.71.6.2496.
DNA of bacteriophage lambda recombines in a cell-free extract prepared from an induced Escherichia coli lysogen of bacteriophage lambda. The assay for recombination in vitro takes advantage of the ability of such an extract to package lambda DNA and to assemble complete phage particles. For example, when lambda DNA that has been extracted from phage with the immunity of 434 is added to an extract, infectious lambda imm 434 particles are produced. The precursor DNA molecule in this packaging reaction is a multichromosomal polymer; circular monomers, for example, are not packaged.Nevertheless, when 434 circular DNA monomers are added to an extract, some phage that contain the imm 434 marker are produced. In this case, the circular DNA had recombined with lambda DNA in the extract and thereby had become part of a polymeric structure, which by the normal packaging process could give rise to infectious particles with the imm 434 marker. Genetic recombination is demonstrated when imm 434 circular monomer DNA carries amber mutations in genes A and B; then most of the 434 plaque formers produced in vitro are A(+)B(+), the genotype of the endogenous lambda DNA. Genetic crossing-over occurs through a region that contains the prophage attachment site, suggesting that recombination is carried out by the lambda Int functions. The 434 recombinant plaque formers are particles physically identical to wild-type 434 particles, as judged by their buoyant density in a CsCl equilibrium gradient.
λ噬菌体的DNA可在从经诱导的λ噬菌体大肠杆菌溶原菌制备的无细胞提取物中发生重组。体外重组检测利用了这种提取物包装λDNA并组装完整噬菌体颗粒的能力。例如，当将从具有434免疫性的噬菌体中提取的λDNA添加到提取物中时，会产生具有感染性的λimm 434颗粒。此包装反应中的前体DNA分子是一种多染色体聚合物；例如，环状单体不会被包装。然而，当将434环状DNA单体添加到提取物中时，会产生一些含有imm 434标记的噬菌体。在这种情况下，环状DNA已在提取物中与λDNA重组，从而成为聚合物结构的一部分，通过正常的包装过程可产生带有imm 434标记的感染性颗粒。当imm 434环状单体DNA在基因A和B中携带琥珀突变时，证明发生了基因重组；然后在体外产生的大多数434噬菌斑形成体是A(+)B(+)，即内源性λDNA的基因型。遗传交换发生在一个包含原噬菌体附着位点的区域，这表明重组是由λ整合酶功能进行的。通过在CsCl平衡梯度中的浮力密度判断，434重组噬菌斑形成体颗粒在物理上与野生型434颗粒相同。
