Construction and characterization of new cloning vehicle. II. A multipurpose cloning system

✍ Scribed by Bolivar, Francisco; Rodriguez, Raymond L.; Greene, Patricia J.; Betlach, Mary C.; Heyneker, Herbert L.; Boyer, Herbert W.; Crosa, Jorge H.; Falkow, Stanley

• Book ID: 120368556
• Publisher: Elsevier Science
• Year: 1977
• Tongue: English
• Weight: 930 KB
• Volume: 2
• Category: Article
• ISSN: 0378-1119
• DOI: 10.1016/0378-1119(77)90000-2

✦ Synopsis

In vitro recombination techniques.were used to construct a new cloning vehicle, pBR322. This plasmid, derived from pBR313, is a relaxed replicating plasmid, does not produce and is sensitive to colicin El, and carries resistance genes to the antibiotics ampicillin (Ap) and tetracycline (Tc). The antibioticresistent genes on pBR322 are not transposable. The vector pBR322 was constxucted in order to have a plasmid with a single PstI site, located in the ampiciUin-resistant gene (Apt), in addition to four unique restriction sites, EcoRI, HindIII, SarnHI and SalI. Survival of Eseherich~a coli stxain X1776 containing pBR313 and pBR322 as a function of thymine and diaminopimelic acid (DAP) starvation and sensitivity to bile salts was found to be equivalent to the non-plasmid containing strain. Conjugal transfer of these plasmicis in bi-and triparental matings were significantly reduced or undetectable relative to the plasmid ColEI.