Doctor of Philosophy (PhD)


Biological Sciences

Document Type



There is mounting evidence of impacts of single base substitutions in both somatic and inherited genetic diseases. However, the occurrence of single base mutations in the germline and gametes has previously only been by conjecture and assumption. There is a need for understanding the risk of offspring inheriting small intragenic mutations, the frequencies at which these mutations are present in germ cells, and the impact that these inherited mutations may have son the predisposition to somatic disease during one’s lifetime. The present study attempts to establish baselines of mutational frequencies in two well characterized genes in human tissues, and demonstrate the prevalence of mutations in two genes, Kristen-ras (KRAS) and TP53 in direct analysis of semen specimens. These two genes play important roles in human disease. Eleven normal human male volunteers’ semen specimens were tested for single base substitutions (SBS). A detection sensitivity of one mutant cell in 106 wild type cells was achieved with the aid of an ultra-sensitive detection method, which utilizes the combined power of the polymerase chain reaction, restriction endonuclease, and ligase chain reaction techniques. The first base of the TP53 gene codon 248, 282, 273 and the second base of codon 273 and the first base of the Kristen-Ras (KRAS) gene codon 12 were studied. The observed mutation prevalence was significantly higher in the first base of the codon 248 of the TP53 gene compared to the first base of codon 273 or the first base of codon 282, both (both P< 0.006). Similarly, there was a significant difference between the prevalence of the first base of codon 12 of the KRAS gene and the first base of codon 273, or the first base of codon 282 (both P< 0.04). Interestingly, the SBS mutation prevalence was nearly the same in the first base of codon 12 KRAS gene and the first base in codon 248 of the TP53 gene (P< 0.006). The spectra of base substitution mutations observed in this study may provide clues to the mutational mechanisms present in human sperm, and may lead to the improved risk assessment for genetic counseling. These data represent the first reports of mutational frequencies in the TP53 and KRAS genes in normal human semen specimens, and demonstrate that single base mutational frequencies for different genes can be obtained directly from semen samples.



Document Availability at the Time of Submission

Release the entire work immediately for access worldwide.

Committee Chair

Vincent L. Wilson