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Nucleic Acids Research, 1978, Vol. 5, No. 7 2381-2390
© 1978

Articles

Screening of cloned recombinant DNA in bacteria by in situ colony hybridization

Brigitte Cami and Philippe Kourilsky*

Groupe de Biologie Moléculaire du Gène, E.R.A. C.N.R.S. No 201, Unité de Géne Génétique, Institut Pasteur 28 Rue du Docteur Roux, 75015 Paris, France

*To whom all correspondence should be addressed

Received April 21, 1978. 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.


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