title: [Cloning of human LUNX gene enhancers and analysis of transcriptional regulation].
abstract: OBJECTIVE
  To identify the enhancers of human lung specific X protein (LUNX) and their regulation at the transcription level in vitro.
  
  
  METHODS
  Three enhancer fragments (E1:+3770~+3959bp; E2: +6454~+6555bp; E3: +14553~+14652 bp) predicted by bioinformatics software were isolated from the human genomic DNA by PCR amplification. Luciferase assay was performed to detect the activities of the enhancers in transcriptional regulation.
  
  
  RESULTS
  PCR products were confirmed by DNA sequencing. The amplified enhancers digested by Kpn I/Xho I and BamH I/Sal I, to generate the sticky-end fragments were inserted into PGL3-promoter in a reporter vector, and 6 luciferase expression vectors were obtained. All the reporter plasmids and pGL3-promoter were transiently transfected into HEK293 cells with an internal control of pSV-β-Galactosidase reporter vector. The enhancer activity of each construct was evaluated by luciferase assay of the cell extracts after transfection for 48 h. The results showed that the 3 fragments, when located upstream, did not increase transcription of reporter gene, but when at the downstream, E1 and E3 increased the transcription by 2.83 and 1.59 folds of that of pGL3-promoter, respectively.
  
  
  CONCLUSIONS
  LUNX gene sequences from +3770 to +3959 bp and +14553 to +14652 bp possess the capacity to enhance gene transcription.
authors.affiliation: Department of Pathology, College of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, China.
authors.forename: Yong-jian
authors.initials: YJ
authors.lastname: Deng
authors.forename: Shuang
authors.initials: S
authors.lastname: Wang
authors.forename: Lin
authors.initials: L
authors.lastname: Zheng
authors.forename: Na
authors.initials: N
authors.lastname: Tang
authors.forename: Xiao-qin
authors.initials: XQ
authors.lastname: Xiao
mesh_terms: D001483:Base Sequence; D003001:Cloning, Molecular; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D006023:Glycoproteins / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000235:genetics*; D014158:Transcription, Genetic
chemical_list: C485599:BPIFA1 protein, human; D006023:Glycoproteins; D010750:Phosphoproteins