Characterization of phosphofructokinase-M gene expression in preimplantation mouse embryos through the use of competitive reverse transcription-polymerase chain reaction

The preimplantation mouse embryo undergoes many metabolic changes as development proceeds. One major change is the switch from a pyruvate based metabolism, to a glucose based metabolism. The phosphofructokinase enzyme is the regulatory enzyme of glycolysis and is thought to be a major contributor in controlling the block to glycolysis in early preimplantation mouse embryos. This study was undertaken to construct a system that would allow detection of RNA for the highly glycolytically active subunit (muscletype) of the phosphofructokinase (PFK) enzyme. A muscle specific mutant PFK plasmid was generated to provide mutant internal control RNA. Using this internal control, initial reverse transcriptionpolymerase chain reaction data collected from early embryo stages suggest that the muscle type PFK subunit RNA is not expressed in the preimplantation mouse at the 1-cell or blastocyst stages. This result suggests that PFK activity detected at the later morula and blastocyst stages must be from either a different PFK subunit or a novel embryonic form of PFK.Department of BiologyThesis (M.S.