Siccna2

Human endometrial cancer cell lines (Ishikawa and AN3CA) were purchased from Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences (Bei Jing, China). Ishikawa, AN3CA were cultured in DMEM (10% fetal bovine serum) at 37 °C, 5% CO2 condition. siCDC20 and siCCNA2 purchased from Guangzhou Ribobio Co., Ltd. and transfected into cells by Lipofectamine RNAiMAX. The target sequence of siCDC20 was 5′-GACCACTCCTAGCAAACCT-3′, the target sequence of siCCNA2 was 5′-GCTGTGAACTACATTGATA-3′. After 48 hours of transfection, western blot was used to detect the efficiency of gene knockdown. The western blot was performed as previously described^{43 (link)}. The primary antibodies were anti-CDC20 (1:1000, Abcam, ab26483), anti-CCNA2 (1:1000, Abcam, ab181591) and anti-β-actin (1:1000, Abcam, USA). After 48 hours of transfection, cell growth curve (Cell Counting Kit-8 assay) was used to evaluate the proliferation of each group of cells. The CCK-8 assay was performed as previously described^{44 (link)}.

We selected prostate cancer cell lines C42 and PC3 for further study. The cell line was purchased from the China Collection Culture Center (Wuhan, China). Both cell lines were cultured in PRMI- 1640 medium containing 10% fetal bovine serum and 1% penicillin-streptomycin, in the incubator containing 37 ℃, 5% nitrogen dioxide. Fetal bovine serum, penicillin-streptomycin, and PRMI-1640 were purchased from Biyuntian Company (shanghai, China). We used siRNA to downregulate the expression of CCNA2. First, C42 and PC3 cells were seeded at a density of 2*10⁵ per well on a six-well plate, and si-CCNA2 and si-mock were inoculated separately when the cells were grown to about 80%. Transfection reagents, si-CCNA2, and si-mock were purchased from Ribobio Biotech Company (Guangzhou, China). The si-mock was regarded as the NC group, and the si-CCNA2 was regarded as the SI group.