In-vitro headful packaging system for cloning dna fragments...

Details In-vitro headful packaging system for cloning dna fragments... In-vitro headful packaging system for cloning dna fragments...

C - Chemistry, Metallurgy

12

N

195/1.12, 195/1.

C12N 15/63 (2006.01) C12N 1/21 (2006.01)

Patent

CA 1337552

This invention relates to an in vitro headful packaging process for cloning foreign DNA fragments as large as 95 kb comprising: modifying vector DNA by inserting a stuffer fragment into a blunt end producing site which is proximal to a pac site; digesting the product of step (a) to produce two vector arms each of which contains (i) a blunt end, (ii) another end which is compatible with the foreign DNA fragment which is to be cloned, and (iii) a loxP site; ligating the foreign DNA to the product of step (b) without generating concatemers; reacting the product of step (c) with pac cleavage proficient extract and head-tail proficient extract wherein the ratio of large heads to small heads in the head-tail extract is at least 5:1; infecting a Cre+ bacterial strain with the product of step (d); and recovering the cloned DNA.

613449

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