Cloning of New Antigen Gene MLAA-34 Promoter and Identification of Core Region in Acute Monocytic Leukemia

Abstract

To clone the promoter sequence of acute monocytic leukemia new antigen gene.MLAA-34 and identify its promoter core region. The full-length fragment of MLAA-34 gene promoter region was amplified by PCR, then was ligated into pGL3-Basic vector, and the recombinant plasmid was cloned. Constructed a series of MLAA-34 gene promoter 5' flanking region truncated plasmid. These recombinant plasmids were transfected into U937 and HEK293 cells, and the dual luciferase reporter gene was used to detect the promoter activity of each fragment to determine the minimum active region. Transcription factor binding sites were analyzed by bioinformatics methods. The recombinant plasmid containing MLAA-34 promoter sequence and its truncated plasmid were successfully constructed, and the promoter activity was significantly increased as compared with the empty vector (P＜0.001). The minimal active region of MLAA-34 located between 402 bp and 200 bp. It contained multiple transcription factor binding sites such as E2F1, MZF-1, SP1, USF2 and STAT3. The promoter of luciferase reporter gene has been successfully constructed with different deletion fragments of MLAA-34, and its core promoter region may contain multiple transcription factor sequence.