Excising Phoa from E. Coli to Study Topologies of Possible Membrane Proteins
Location
CSU Ballroom
Start Date
9-4-2012 10:00 AM
End Date
9-4-2012 11:30 AM
Student's Major
Biological Sciences
Student's College
Science, Engineering and Technology
Mentor's Name
Timothy Secott
Mentor's Department
Biological Sciences
Mentor's College
Science, Engineering and Technology
Description
One way to study the topology, or shape, of a protein in a cell membrane is to fuse the gene encoding the protein of interest with another gene encoding a protein that has the ability to indicate the shape of the protein of interest in the membrane. The alkaline phosphatase protein, encoded by the phoA gene, has this ability and is therefore useful in studying membrane protein topologies in E. coli. However, E. coli contain the phoA gene in their genome, making it possible for the phoA gene in the fusion construct to recombine into the genome and render the fusion construct useless. To circumvent this issue, we intend to excise the phoA gene from the genome of a strain of E. coli. Thus far, we have generated a short segment of DNA that has ends similar to the ends of the phoA gene in E coli’s genome. This segment is currently being used to displace the phoA gene from E. coli’s genome. Following this process the segment will be removed from the genome, leaving a strain of E. coli lacking the phoA gene. We intend to utilize this strain and a phoA fusion construct to study the topology of the mammalian cell entry protein of Mycobacterium avium subspecies paratuberculosis, which is thought to help initiate a damaging gastrointestinal disease in cattle. In addition, generating this strain of E. coli will provide investigators at Minnesota State University, Mankato with the ability to study the topologies of other important membrane proteins.
Excising Phoa from E. Coli to Study Topologies of Possible Membrane Proteins
CSU Ballroom
One way to study the topology, or shape, of a protein in a cell membrane is to fuse the gene encoding the protein of interest with another gene encoding a protein that has the ability to indicate the shape of the protein of interest in the membrane. The alkaline phosphatase protein, encoded by the phoA gene, has this ability and is therefore useful in studying membrane protein topologies in E. coli. However, E. coli contain the phoA gene in their genome, making it possible for the phoA gene in the fusion construct to recombine into the genome and render the fusion construct useless. To circumvent this issue, we intend to excise the phoA gene from the genome of a strain of E. coli. Thus far, we have generated a short segment of DNA that has ends similar to the ends of the phoA gene in E coli’s genome. This segment is currently being used to displace the phoA gene from E. coli’s genome. Following this process the segment will be removed from the genome, leaving a strain of E. coli lacking the phoA gene. We intend to utilize this strain and a phoA fusion construct to study the topology of the mammalian cell entry protein of Mycobacterium avium subspecies paratuberculosis, which is thought to help initiate a damaging gastrointestinal disease in cattle. In addition, generating this strain of E. coli will provide investigators at Minnesota State University, Mankato with the ability to study the topologies of other important membrane proteins.
Recommended Citation
Scheid, Adam. "Excising Phoa from E. Coli to Study Topologies of Possible Membrane Proteins." Undergraduate Research Symposium, Mankato, MN, April 9, 2012.
https://cornerstone.lib.mnsu.edu/urs/2012/poster-session-A/18
