Molecular Genetics in Rhodopseudomonas Sphaeroides: Transformation, Plasmids, and Gene Cloning

A Tris-dependent transformation system was developed for R. sphaeroides. Washing the cells with 0.5M Tris-buffer, pH 7.2, at 0(DEGREES)C prior to exposure to DNA was found to be necessary for inducing a competent state for DNA uptake. Polyethylene glycol (PEG) and Ca('2+) were also determined to be necessary components of this system and were added at the time of DNA addition to Tris-treated cells. The transformation method was most effective with plasmid DNA in the CCC configuration and frequencies as high as 1 x 10('-5) transformants per viable cell were achieved using RSF1010 DNA, although other plasmids, such as pRK290 and RP4 were capable of transforming Tris-treated cells. A versatile cloning vector, pUI81, was constructed from RSF1010 and a pBR322 derivative, pSL25 and shown to be effective in transforming Tris-treated R. sphaeroides.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Microbiology

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

1983

Author dc:creator

• Fornari, Chester Stephen

• Biology, Microbiology

Identifier

(UMI)AAI8409923

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/71163