[Comparison and analysis of human dentin matrix protein 1 promoter activity in three different cells]
Jian-Liang Pang1, Bu-Ling Wu, Ya-Qing Zhang
1Dept. of Operative Dentistry and Endodontics, College of Stomatology, The Fourth Military Medical University, Xi'an 710032, China.
Objective:
To observe and compare the luciferase activities of different length segments of human dentin matrix protein 1 promoter in human dental pulp stem cells (HDPSC), osteoblasts (OC) and Hela cells.
Methods:
The differentlength desired DNA segments were obtained from 2 195 bp Dmp1 promoter cloned by PCR method. The amplified promoter segments with different length were cloned into luciferase report gene vector pGL3-Basic, the correct orientation of those inserts was verified by cutting with two different restrict enzymes. The luciferase activity was observed after different pGL3-PDmp1 vectors were transfected transiently into those three different-type cells.
Results:
6 Dmp1 promoter segments with different-length were obtained successfully, and luciferase report gene vectors with different promoter segments were successfully constructed after identified by restriction enzymes cutting. They had different luciferase activities when they were transfected transiently into HDPSC, and the region of -505(-)-193 bp and -935(-)-505 bp could be regarded as the specific promoters of Dmp1 promoter for HDPSC and OC respectively, which could include the basic regulatory elements.
Conclusion:
The correct clone of the upstream of human Dmp1 promoter segments with different length had been obtained, and they had strong luciferase activities in HDPSC and OC, but very low in Hela cell. These results will make an important basis for studying mineralized tissue-specific transcriptional regulation mechanisms of Dmp1.
