VIEW ARTICLE

Genetics

Transformation of Erwinia herbicola with Plasmid pBR322 Deoxyribonucleic Acid. George H. Lacy, Assistant plant pathologist, Department of Plant Pathology and Botany, The Connecticut Agricultural Experiment Station, New Haven, CT 06504; Robert B. Sparks, Jr., assistant geneticist, Department of Genetics, The Connecticut Agricultural Experiment Station, New Haven, CT 06504. Phytopathology 69:1293-1297. Accepted for publication 13 June 1979. Copyright 1979 The American Phytopathological Society. DOI: 10.1094/Phyto-69-1293.

Erwinia herbicola was transformed by a CaCl₂ technique with plasmid pBR322 deoxyribonucleic acid (DNA) at frequencies up to 1 × 10^–6 transformants per recipient cell (TPR). Since pBR322 carries resistance to ampicillin (Ap^r) and tetracycline (Tc^r), transformants were selected on media containing either one or both of those antibiotics. Covalently closed circular (CCC) plasmid DNA was isolated from lysates of E. herbicola (pBR322) transformants by equilibrium centrifugation in cesium chloride-ethidium bromide gradients. The CCC DNA of E. herbicola (pBR322) was found by electron microscopy and gel electrophoresis to contain a DNA species the same size as that of pBR322. This plasmid had the same restriction pattern as pBR322. Transformation of Escherichica coli with CCC DNA from E. herbicola/ pBR322 yielded Ap^r Tc^r transformants at frequencies as high as 1 × 10^–4 TPR. The CCC DNA isolated from all E. coli transformants tested had electrophoresis patterns identical to those of pBR322. Transformation of pBR322, a recombinant DNA cloning vehicle, into E. herbicola indicates that development of a gene cloning system in this species may be possible.