Establishment　and identification of an ovarian cancer cell strain stably expressing CDC50A

Abstract

Objective To establish a stable-expressing human cell cycle regulatory protein 50A (CDC50A) SKOV3 cell strain expressing vector containing CDC50A by molecular cloning. Methods CDC50A was introduced into the expression vector pLVX-IRES-GFP by molecular cloning method. The pLVX-CDC50A-GFP expression plasmid was packaged by lentivirus and stably transfected into SKOV3. SKOV3 cell strain with high-expressing CDC50A was established. The expression of Flag protein was analyzed by flow cytometry, Western blot and immunofluorescence. Results The construction of pLVX-CDC50A-GFP expressing vector was confirmed by gel electrophoresis and endonuclease digestion. The efficiency of infection was observed qualitatively under fluorescence microscope and detected quantitatively by flow cytometry as 86.5%. Flag-tag protein was detected by Western blot to present only in the positive group. Conclusions The SKOV3 cell strain was established by infecting with lentivirus of pLVX-CDC50A-GFP, laying a foundation for the mechanism study of CDC50A in ovarian cancer.

Keywords

• cdc50a|over-express|ovarian cancer|stable expression