Mouse Spermatogenesis At Different Stages Of Spermatogenic Cell Gene Expression Profile

Spermatogenesis is a unique cellular differentiation process during which diploid spermatogonia differentiate to haploid round spermatids after two meiotic divisions. Subsequently, the round spermatids develop into elongating spermatids undergoing HPRR (histone-protamine replacement reaction), nucleus condensation, and morphological changes. So far, the molecular mechanisms of spermatogenesis remain unclear. Only a few of key genes controlling spermatogenesis were reported, although more and more spermatogenesis-related genes were found. In the present study, newly developed techniques such as fluorescent DDRT-PCR and cDNA microarray were used to analyze the expression characteristics of numerous genes in different stages of spermatogenic cells. A number of differentially expressed genes in spermatogenesis were found. Further studies on the regulation and function of these differentially expressed genes during spermatogenesis may provide insight into molecular mechanisms of spermatogenesis and be helpful for identification of the key genes to regulate spermatogenesis. The present thesis includes three parts following.1. Isolation of different types of spermatogenic cells. Using the method of velocity sedimentation at unit gravity followed by cell attachment incubation, we obtained 94% purity of primitive type A spermatogonia, 90% purity of type B spermatogonia, 88% purity of preleptotene spermatocytes, 95% purity of pachytene spermatocytes , 96% purity of round spermatids and 92% purity of elongating spermatids from testis of 6, 9, 14, 21, 35 and 60 days old mice, respectively. We also get 98% purity of mature spermatozoon from epididymis of 12 weeks old of mice by centrifugation.2. Cloning of ESTs of spermatogenesis-related genes from the early stages of spermatogenic cells of mice using a modified DDRT-PCR method. The fluorescent differential display technique was performed, instead of radioactive differential display, to screen differentially expressed genes between primitive type A spermatogonia and type B spermatogonia. Differences in expression of the...