Linked Life Data

8938437

Source:http://linkedlifedata.com/resource/pubmed/id/8938437

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pubmed-article:8938437pubmed:abstractTextThis study addresses two important technical problems: how to perform targeted alterations such as site-directed mutagenesis and deletions in large fragments of DNA and how to construct full-length genes from two partly overlapping bacterial artificial chromosome (BAC) plasmids. Given the size and the lack of convenient unique restriction sites in these large-insert bacterial clones, these are nontrivial tasks. Here we describe a simple and efficient protocol based on RecA-assisted restriction endonuclease (RARE) cleavage, a method that enables sequence-specific cleavage of genomic DNA. The same protocol has been used with minor modifications to introduce site-specific mutations into an apolipoprotein-B 90-kb P1 clone, to generate deletions in a 160-kb BAC, and to generate a 160-kb BAC containing the complete 92-kb gene for low-density lipoprotein-related protein-1 (LRP-1) from two smaller overlapping BACs ("BAC marriage").lld:pubmed
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pubmed-article:8938437pubmed:dateRevised2007-11-14lld:pubmed
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pubmed-article:8938437pubmed:articleTitleA simple and efficient method for making site-directed mutants, deletions, and fusions of large DNA such as P1 and BAC clones.lld:pubmed
pubmed-article:8938437pubmed:affiliationGladstone Institute of Cardiovascular Disease, University of California, San Francisco 94141-9100, USA. jan-boren.gicd@quickmail.ucsf.edulld:pubmed
pubmed-article:8938437pubmed:publicationTypeJournal Articlelld:pubmed
pubmed-article:8938437pubmed:publicationTypeResearch Support, U.S. Gov't, P.H.S.lld:pubmed
pubmed-article:8938437pubmed:publicationTypeResearch Support, Non-U.S. Gov'tlld:pubmed
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