title: Construction of shRNA-Bmi-1 plasmid and its effect on proliferation of ECA109 cells.
abstract: OBJECTIVE
  To explore the role of Bmi-1 gene in the proliferation of squamous carcinoma cells and whether the silencing Bmi-1 can inhibit the growth of squamous cell carcinomas cells.
  
  
  METHODS
  The expressions of Bmi-1 in primary cultured Fibroblasts, karatinocyte cell line Hacat,squamous carcinoma cell line A431, and ECA109 were detected by reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis. Recombinant plasmid inserted with Bmi-1 gene short hairpin RNA (shRNA) expression vector PGPU6/GFP/Neo-shBmi-1 was constructed and transfected into ECA109 cells with control set. After transfection for 48 and 72 hours,the mRNA and protein levels of Bmi-1 were examined with RT-PCR and Western blot analysis, respectively. The proliferation of the ECA109 cells was evaluated by MTT method and flow cytometry.
  
  
  RESULTS
  Bmi-1 was highly expressed in A431 and ECA109 cells than in Fibroblast cells and Hacat cells. The mRNA and protein expressions of Bmi-1 were significantly silenced in ECA109 cells after recombinant expression vector PGPU6/GFP/Neo-shBmi-1 transfection (P<0.05). Compared with the control groups,the proliferation of ECA109 transfected with PGPU6/GFP/Neo-shBmi-1 was significantly inhibited (P<0.05), and cells in G1 phase increased while in S phase decreased (P<0.05).
  
  
  CONCLUSIONS
  Bmi-1 is involved in the proliferation of squamous carcinoma cells. After the silencing of Bmi-1 expression,the proliferation ECA109 cells is suppressed due to the altered cell cycle.
authors: Department of Dermatology, the Second Affiliated Hospital, Xi'an Jiaotong University, Xi'an 710004, China.
authors: Ming
authors: M
authors: Zhou
authors: Department of Dermatology, the Second Affiliated Hospital, Xi'an Jiaotong University, Xi'an 710004, China.
authors: Song-mei
authors: SM
authors: Geng
mesh_terms: D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell; D002453:Cell Cycle; D002460:Cell Line; D049109:Cell Proliferation; D005347:Fibroblasts; D005434:Flow Cytometry; D006801:Humans; D010957:Plasmids; D063150:Polycomb Repressive Complex 1; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014162:Transfection
chemical_list: C069077:BMI1 protein, human; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D063150:Polycomb Repressive Complex 1