Acquisition of Quantitative Instrumentation for Research in Molecular Biology and Biochemistry

Timothy Finco and Douglas Fantz, Departments of Biology and Chemistry, Agnes Scott College.

INTRODUCTIONIn the Fall of 2009, Dr. Finco of the Biology Department and Dr. Fantz of the Chemistry Department were awarded a grant from the National Science Foundation (NSF-MRI) to purchase a set of four instruments to promote research by undergraduates in the fields of biochemistry and molecular biology. This poster will highlight the uses of these instruments as tools to conduct quantitative analysis of biomolecules in both the research and teaching laboratories at Agnes Scott. In addition, Drs. Fantz and Finco will schedule a time to demonstrate these instruments to interested students in room #204E of the Bullock Science Center.

This instrument allows us to monitor the progress of a PCR reaction in order to quantify the amount of template DNA in a sample. This will allow us to study gene expression by measuring the amount of mRNA transcripts in cells. Dr. Finco will be using this instrument to quantify the expression of the LAT gene in cultured cells to understand LAT’s regulation during the development of the immune system.

Phosphorimager: BIO-RAD Personal Molecular Imager System

This instrument allows for the imaging of molecules that are fluorescent or chemiluminescent. Often nucleic acids of different sizes such as DNA or RNA can be separated on an agarose gel and treated with a fluorescent chemical such as ethidium bromide for visualization. Another application of this instrument is the imaging and quantification of proteins through Western Blotting, a technique that utilizes enzyme-linked antibodies that are able to catalyze light-emitting reactions.

This instrument allows for the precise quantification of light energy. Molecular signal transduction pathways can often be monitored in cells by coupling the activity of a molecule to the light-generating enzyme, luciferase. This instrument allows for the quantification of the firefly and sea pansy luciferase enzymes. Drs. Fantz and Finco will use this instrument to quantify transcription factor activity and promoter element function.

The purchase of these instruments was funded by the National Science Foundation’s Major Research Instrumentation Grant (MRI-R2) #0959416. Computers used to interface with this instrumentation were provided by this grant as well as the Agnes Scott Department of Biology. We would also like to acknowledge Nancy Devino, and Sherri Williams for assistance in the preparation and submission of the grant, and Tamara Carter for assistance in instrument setup.

Real-Time PCR: BIO-RAD CFX96 System

Molecular Imager: BIO-RAD Chemi-Doc XRS+ System

Luminometer: Promega GloMax96 System

Acknowledgements

Often biological molecules such as DNA and proteins must be radioactively labeled in order to be detected. This instrument allows us to quantify radioactive biomolecules without the use of film. Radioactive samples are exposed to an imaging plate and the energy emitted by the sample is stored in phosphor crystals. A laser then scans the screen and the stored energy can be efficiently quantified. Dr. Fantz will be using this instrument to detect the binding of a transcription factor protein to a DNA target.