Abstract: AIM: To construct a lentiviral vector of RNA interference (RNAi) of ERα gene and transfect them into HEK-293T cells to detect the titer of virus. METHODS: The effective sequence of siRNA targeting ERα gene was confirmed in our previous study. The complementary DNA containing both sense and antisense Oligo DNA of the targeting sequence was designed, synthesized and cloned into the pGCSIL-GFP vector, which contained U6 promoter and green fluorescent protein (GFP). The resulting lentiviral vector containing ERα?shRNA was named shERα-LV, and it was confirmed by PCR and sequencing. HEK-293T cells were cotransfected with lentiviral vector shERα-LV，pHelper 1.0 and pHelper 2.0. All virus stocks were produced by LipofectamineTM 2000 transfection. The titer of virus was tested according to the expression level of GFP. CONCLUSION: PCR and DNA sequencing demonstrated that shERα-LV producing ERα-shRNA was constructed successfully. The titer of concentrated virus was 2E+8TU/ml. DISCUSSION: The lentivirus RNAi vector of ERα?was constructed successfully. Our work provides the basis for ERα signal transduction pathway in breast cancer.

Key words: RNA interference, ERα, Plasmid construction, Lentivirus