E.i. Dupont De Nemours and Company

Nat L Sternberg

Brian L Sauer

James C Pierce

This relates to an in vitro headful packaging system for cloning foreign DNA fragments as large as 95 kb. Also of concern is a method of cloning and controlling amplification of DNA fragments as large as 95 kb in a vector containing a multicopy replicon under control of a lac promoter as well as vectors useful for cloning foreign fragments of DNA as large as 95 kb and lysogens to prepare pac-cleavage proficient extract and head-tail proficient extract.

Vitro headful packaging system for cloning DNA fragments as large as 95kb

Positive selection cassettes are disclosed which contain a lethal gene, a promoter, a repressor sequence overlapping the promoter, and a cloning site between the promoter and the lethal gene. Insertion of a foreign nucleic acid sequence into the cloning site prevents expression of the lethal gene. Expression of the lethal gene under nonrepressed conditions kills a host organism containing a positive selection cassette which does not contain the foreign nucleic acid sequence.

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West Chester, PA, United States of America

Nat L Sternberg