Cami B, Kourilsky P
Nucleic Acids Res. 1978 Jul;5(7):2381-90. doi: 10.1093/nar/5.7.2381.
We have developped in situ methods of colony hybridization in which there is no need to replicate colonies one by one prior to hybridization. The best method consists in promoting partial lysis of the colonies on the plates by means of a resident thermoinducible prophage. It appears that colonies are heterogeneous with respect to prophage induction, so that survivors remain in each colony. Blotting onto nitrocellulose filters and hybridization with a highly radioactive probe permits the screening of many thousands of colonies per plate for the presence of a DNA sequence carried by a plasmid and complementary to the probe. This procedure greatly facilitates the isolation of recombinant plasmids which carry a specific DNA sequence. We also describe a second, less efficient procedure which does not use prophage induced lysis, and is potentially usable with B2 or EK2 safety systems, without modification of the bacterial hosts.
我们已经开发出原位菌落杂交方法，在杂交前无需逐个复制菌落。最佳方法是利用驻留的热诱导原噬菌体促使平板上的菌落部分裂解。似乎菌落对于原噬菌体诱导是异质的，这样每个菌落中都会有存活者。将其印迹到硝酸纤维素滤膜上并与高放射性探针杂交，可对每块平板上数以千计的菌落进行筛选，以检测由质粒携带且与探针互补的DNA序列的存在。该程序极大地促进了携带特定DNA序列的重组质粒的分离。我们还描述了第二种效率较低的程序，该程序不使用原噬菌体诱导裂解，并且在不修饰细菌宿主的情况下，可能适用于B2或EK2安全系统。
