Effect Of NOB1 Gene Silence On Cell Proliferation And Cell Cycle Progression In Human Ovarian Cancer SKOV3 Cell

Objective:We study the growth of ovarian cancer SKOV3 cells in vitro propagation, cell cycle and apoptosis of human NOB1 gene by siRNA interference, a preliminary study of NOB1 gene in the role in the development of ovarian cancer, for the study of molecular regulation mechanism in ovarian cancer and other tumors in the occurrence, development process, and provides new clues and reference for the study of molecular mechanism and human NOB1 gene as a target for cancer treatment.Method:1. Culture the human ovarian cancer cell line SKOV3 to the log phase, then divided into the 6-well-plate.2. Using the Santa Cruz si RNA Transfection Reagent to transfect Control siRNA(Flourescein Conjugate)-A into human ovarian cancer SKOV3 cells, then observe the transfection efficiency.3. The cell was divided into blank control group、transfection reagent control group、interference group and negative control group, Cruz siRNA Transfection Reagent transfection reagent as the carrier of the negative control Control siRNA-A and interference fragment of NOB1 P siRNA(h) were transfected into human ovarian cancer SKOV3 cells, and then observed the cell under the morphology and calculate growth status.4. Observe the proliferation of cells by the MTT method.5. Using the RT-PCR to observe NOB1 mRNA after transfection.6. To extract the total protein in each group after transfection, expression of NOB1 protein of the cells were detected by Western blotting method.7. Using the flow cytometry to check the cell apoptosis and cell cycle.Results:1. The Control siRNA(Flourescein Conjugate)-A was transfected into the human ovarian cancer SKOV3 cells, placed under the inverted fluorescence phase contrast microscope to observe, we found that most of the human ovarian cancer SKOV3 cells were transfected in green fluorescent, and the coverage of the green fluorescence was more than 80%.2.The MTT results showed that the proliferation of the NOB1 interference group was decreased obviously.3, The RT-PCR detected NOB1 gene mRNA, the expression of NOB1 in the interference group was significantly decreased than the other 3 groups,(P <0.05).4.Western blotting detected that the NOB1 protein expression in the NOB1 interference group was significantly decreased than the other 3 groups(P <0.05).5. Flow cytometry apoptosis results show NOB1 interference cell apoptosis rate increased significantly in group(P <0.05); detection of cell cycle of different group showed, in the NOB1 interference group the number of G1 cells was more than the other three groups, indicating that the cells in the NOB1 interference group arrest in the G1 phase, and they were difficult to enter the S phase.Conclusion:The interference fragment of NOB1 P siRNA(H) can be transfected into SKOV3, the NOB1 P siRNA(H) can inhibit the expression of NOB1 in SKOV3, and it can inhibit the proliferation and promote apoptosis in SKOV3.