CMOP Undergraduate Intern Mentoring Opportunity
Deadline: March 26, 2010
Selections Announced: April 2, 2010
Name/Title/Institution(s) of senior mentor(s): Brad Tebo, EBS Division Head, EBS
Name/Title/Institution(s) of frontline mentor(s): Kati Geszvain, Senior Research Associate, EBS
Project Title: Regulation of Mn oxidation in Pseudomonas putida GB-1.
Context for Project:
Bacterial manganese (II) oxidation has a profound impact on biogeochemical cycling of Mn, as well as on
the availability of the trace metals adsorbed to the surfaces of solid Mn (III, IV) oxides. To begin to
understand at the molecular level how and why bacteria oxidize Mn, the Tebo lab employs the genetically
tractable Mn oxidizing bacterium Pseudomonas putida GB-1.
Brief Description.
Previous work in the Tebo group suggests that the Mnx two-component regulatory (TCR) pathway is
essential for Mn(II) oxidation. This project will focus on the three genes that comprise this pathway,
employing random mutagenesis and screening to identify constitutively active alleles of these genes. We
will also generate specific mutations in the response regulator component of this pathway, MnxR, which
would be predicted to make this regulator constitutively active. In addition to the Mnx pathway, we will
also investigate the role of other regulators of Mn(II) oxidation, such as RsmA, through generating
plasmids bearing the regulator genes.
Proposed Outcomes/Broader Impact:
TCR pathways sense and respond to changes in the bacterial environment, frequently through altering
expression of downstream genes. In the case of the Mnx, the genes regulated by the pathway may include
those encoding the Mn(II) oxidase enzyme. By generating constitutively active alleles of the Mnx TCR
genes, we hope to produce P. putida GB-1 strains that constitutively express the Mn(II) oxidase. This
would be a valuable tool for the lab for biochemical isolation of the oxidase activity as well as for microarray analysis of the genes co-regulated with the Mn(II) oxidase.
Proposed timeline (within a 10 week span):
Pilot experiments to test screening conditions – 1 week
Random mutagenesis and screening – 3 weeks
Sub-cloning and mapping mutations – 3 weeks
Site-specific mutagenesis and gene cloning – 3 weeks
Intern academic experience and skill set should include:
I would prefer to work with a biology major that has an emphasis on Genetics or molecular biology. This
project is fairly flexible, with approaches that can be tailored to the experience of the student; therefore I
would be willing to work with a younger student.