Presentation Title

Creating and Characterizing an mntH knockout mutant in E. coli

Faculty Mentor

Brianna Cadena, Desiree Duran, Jessica Del Castillo

Start Date

18-11-2017 2:15 PM

End Date

18-11-2017 3:15 PM

Location

BSC-Ursa Minor 64

Session

Poster 3

Type of Presentation

Poster

Subject Area

biological_agricultural_sciences

Abstract

Under excessive amounts of hydrogen peroxide a cell will activate genes to protect itself against oxidation. In Escherichia coli, one of these genes, mntH, codes for a manganese import protein. Manganese is a redox cofactor and is thought to help protect proteins from oxidative damage. We wish to comprehend the role manganese may have in DNA damage and repair. To study this, we have successfully knocked out the mntH gene in a reporter strain of E. coli which is susceptible to DNA mutation. We used the Lambda Red system, a special recombination system, to insert a linear piece of DNA containing the kanamycin gene into the E. coli's genomic DNA, replacing the mntH gene. To optimize our technique, we successfully performed the knockout in a commercially available E. coli strain DH5-α, known to efficiently accept foreign DNA. We managed to generate a growth curve of DH5-alpha of the wild type and knockout mutant. Our results indicate that the mntH knockout grows more slowly than the wild type in the presence of hydrogen peroxide, which supports our hypothesis that mntH protects E. coli against oxidative stress.

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Nov 18th, 2:15 PM Nov 18th, 3:15 PM

Creating and Characterizing an mntH knockout mutant in E. coli

BSC-Ursa Minor 64

Under excessive amounts of hydrogen peroxide a cell will activate genes to protect itself against oxidation. In Escherichia coli, one of these genes, mntH, codes for a manganese import protein. Manganese is a redox cofactor and is thought to help protect proteins from oxidative damage. We wish to comprehend the role manganese may have in DNA damage and repair. To study this, we have successfully knocked out the mntH gene in a reporter strain of E. coli which is susceptible to DNA mutation. We used the Lambda Red system, a special recombination system, to insert a linear piece of DNA containing the kanamycin gene into the E. coli's genomic DNA, replacing the mntH gene. To optimize our technique, we successfully performed the knockout in a commercially available E. coli strain DH5-α, known to efficiently accept foreign DNA. We managed to generate a growth curve of DH5-alpha of the wild type and knockout mutant. Our results indicate that the mntH knockout grows more slowly than the wild type in the presence of hydrogen peroxide, which supports our hypothesis that mntH protects E. coli against oxidative stress.