Developmental expression of GCNF during organ maturity

Thesis (M.S.)--Wichita State University, College of Liberal Arts and Sciences, Dept. of Biological SciencesWithout a ligand, Germ Cell Nuclear Factor (GCNF) binds to and represses genes. Targets include Oct4 (involved in the pluripotency of embryonic and germ cells), CRIPTO-1 (found in undifferentiated stem cells), and reproductive genes in both the male and female including GDF-9 and BMP-15. Previous studies using qRtPCR suggest that GCNF is involved in cell growth proliferation. It was theorized that higher concentrations of GCNF would be found in growing tissues, for instance in embryonic and neonatal tissues, and lower concentrations in fully developed tissues. Tests were conducted by isolating total RNA from hamster tissue samples, reverse transcribing it, and then followed up by RtPCR. Standard curves from isolated ds DNA GCNF PCR product were created to allow the formation of a database monitoring changes in hamster tissue GCNF mRNA levels using ds DNA PCR product. Linearity of the GCNF qRtPCR was reproducible over a broad range of DNA levels. However, it was found that the sample tissues contained extremely low amounts of GCNF mRNA that could not be measured accurately or reproduced easily even by qRtPCR. GCNF 351 primers were found to work better than the GCNFRT primers due to possible alternate splicing occurring in the mRNA not allowing GCNRT primers to bind. The overall conclusion does not support the hypothesis that GNCF mRNA expression is greater in growing versus non-growing adult tissues