Cloning and regulatory expression of the transcriptional regulatory sequences in human breast cancer related DF3 antigen / 中国普通外科杂志

ABSTRACT

Objective To clone the transcriptional regulatory sequences (TRS) in human breast cancer related DF3 antigen, and to test the relationship between the activity of the TRS and the cell surface DF3 antigen. Methods Authors designed a pair of primers according to the registered 5′-flanking region of DF3 antigen. The 771 base pairs of DNA fragment were amplified from the genomic DNA of human MCF-7 breast carcinoma cells by PCR, and cloned to the pMD18-T vector. The results were tested by restrictive enzyme analysis and DNA sequencing. The DF3 TRS was cut by double enzyme Mlu I、Hind III,and cloned to the pGL3 vector . The activity of the DF3 TRS was expressed by analyzing the relative luciferase activities. Results Restrictive endonuclease identification and DNA sequencing proved that the sequence authors got, was correct. The luciferase activity in MDA-MB-231 was hardly detected, whereas in MCF-7 the luciferase activity was about 200 times than in MDA-MB-231. Conclusions The DF3 TRS was cloned successfully. The DF3 activity has a distinct relationship with DF3 antigen. The study shows that DF3 TRS can be used in the gene therapy of breast carcinoma.