| 摘要 |
<p>This invention relates to an in vitro headful packaging system for cloning foreign DNA fragments as large as 95kb comprising: (a) modifying vector DNA by inserting a stuffer fragment into a blunt end producing site which is proximal to a pac site; (b) 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; (c) ligating the foreign DNA to the product of step (b) without generating concatemers; (d) 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; (e) infecting a Cre+ bacterial strain with the product of step (d); and (f) recovering the cloned DNA. This invention also concerns vector bacterial strains useful for cloning foreign fragments of DNA as large as 95 kb.</p> |
