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Title: Cloning systems for Rhodococcus and related bacteria

Abstract

A plasmid transformation system for Rhodococcus was developed using an Escherichia coli-Rhodococcus shuttle plasmid. Rhodococcus sp. H13-A contains three cryptic indigenous plasmids, designated pMVS100, pMVS200 and pMVS300, of 75, 19.5 and 13.4 kilobases (Kb), respectively. A 3.8 Kb restriction fragment of pMVS300 was cloned into pIJ30, a 6.3 Kb pBR322 derivative, containing the E. coli origin of replication (ori) and ampicillin resistance determinant (bla) as well as a Streptomyces gene for thiostrepton resistance, tsr. The resulting 10.1 Kb recombinant plasmid, designated pMVS301, was isolated from E. coli DH1 (pMVS301) and transformed into Rhodococcus sp. AS-50, a derivative of strain H13-A, by polyethylene glycol-assisted transformation of Rhodococcus protoplasts and selection for thiostrepton-resistant transformants. This strain was deposited with the ATCC on Feb. 1, 1988 and assigned ATCC 53719. The plasmid contains the Rhodococcus origin of replication. The plasmid and derivatives thereof can therefore be used to introduce nucleic acid sequences to and from Rhodococcus for subsequent expression and translation into protein. The isolated origin of replication can also be used in the construction of new vectors.

Inventors:
 [1];  [2]
  1. (Athens, GA)
  2. (Decatur, GA)
Issue Date:
Research Org.:
University of Georgia Research Foundation
Sponsoring Org.:
USDOE
OSTI Identifier:
867507
Patent Number(s):
4952500
Application Number:
07/151,319
Assignee:
University of Georgia Research Foundation, Inc. (Athens, GA) SRO
DOE Contract Number:  
AS09-80ER10683; FG09-86ER13588
Resource Type:
Patent
Country of Publication:
United States
Language:
English
Subject:
cloning; systems; rhodococcus; related; bacteria; plasmid; transformation; developed; escherichia; coli-rhodococcus; shuttle; sp; h13-a; contains; cryptic; indigenous; plasmids; designated; pmvs100; pmvs200; pmvs300; 75; 19; 13; kilobases; kb; respectively; restriction; fragment; cloned; pij30; pbr322; derivative; containing; coli; origin; replication; ori; ampicillin; resistance; determinant; bla; streptomyces; thiostrepton; tsr; resulting; 10; recombinant; pmvs301; isolated; dh1; transformed; as-50; strain; polyethylene; glycol-assisted; protoplasts; selection; thiostrepton-resistant; transformants; deposited; atcc; feb; 1988; assigned; 53719; derivatives; introduce; nucleic; acid; sequences; subsequent; expression; translation; protein; construction; vectors; ethylene glycol; polyethylene glycol; acid sequence; nucleic acid; acid sequences; escherichia coli; subsequent expression; recombinant plasmid; restriction fragment; plasmid contains; /435/

Citation Formats

Finnerty, William R., and Singer, Mary E. Cloning systems for Rhodococcus and related bacteria. United States: N. p., 1990. Web.
Finnerty, William R., & Singer, Mary E. Cloning systems for Rhodococcus and related bacteria. United States.
Finnerty, William R., and Singer, Mary E. Mon . "Cloning systems for Rhodococcus and related bacteria". United States. https://www.osti.gov/servlets/purl/867507.
@article{osti_867507,
title = {Cloning systems for Rhodococcus and related bacteria},
author = {Finnerty, William R. and Singer, Mary E.},
abstractNote = {A plasmid transformation system for Rhodococcus was developed using an Escherichia coli-Rhodococcus shuttle plasmid. Rhodococcus sp. H13-A contains three cryptic indigenous plasmids, designated pMVS100, pMVS200 and pMVS300, of 75, 19.5 and 13.4 kilobases (Kb), respectively. A 3.8 Kb restriction fragment of pMVS300 was cloned into pIJ30, a 6.3 Kb pBR322 derivative, containing the E. coli origin of replication (ori) and ampicillin resistance determinant (bla) as well as a Streptomyces gene for thiostrepton resistance, tsr. The resulting 10.1 Kb recombinant plasmid, designated pMVS301, was isolated from E. coli DH1 (pMVS301) and transformed into Rhodococcus sp. AS-50, a derivative of strain H13-A, by polyethylene glycol-assisted transformation of Rhodococcus protoplasts and selection for thiostrepton-resistant transformants. This strain was deposited with the ATCC on Feb. 1, 1988 and assigned ATCC 53719. The plasmid contains the Rhodococcus origin of replication. The plasmid and derivatives thereof can therefore be used to introduce nucleic acid sequences to and from Rhodococcus for subsequent expression and translation into protein. The isolated origin of replication can also be used in the construction of new vectors.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {1990},
month = {1}
}

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