title: [Construction of a miR-23a-27a cluster expression plasmid: a preliminary study of its function].
abstract: OBJECTIVE
  To construct a miR-23a-27a cluster expression plasmid and to explore the target genes and function of the cluster.
  
  
  METHODS
  The pre-miR-23a-27a-pcDNA3.1, pre-miR-23a and pre-miR-27a plasmids were cloned by molecular biology method, and their expression efficiency was tested by dual luciferase reporter gene assay and real-time PCR. Several possible target genes of miR-23a and miR-27a were chosen using softwares and further tested by dual luciferase reporter gene assay. Finally, the function of miR-27a was analyzed in MCF-7 cell by Western blot and real-time PCR.
  
  
  RESULTS
  miR-23a and miR-27a were transcribed from pre-miR-23a-27a-pcDNA3.1, pre-miR-23a and pre-miR-27a plasmids in HEK293T cells, and both influenced the MRE of Sprouty2 gene in pRL-TK vector, and only miR-27a influenced the 3'-untranslated regions (UTR) full length of Sprouty2 gene while miR-27a did not influence the 3'-UTR of Sprouty2 gene with the sited-mutation in the MRE. The protein expression level of Sprouty2 gene was altered after transfection of pre-miR-27a-pcDNA3.1 plasmid while the RNA level remained unchanged.
  
  
  CONCLUSIONS
  Sprouty2 may be the functional target gene of miR-27a, and the construction of plasmids in the study may provide a fundamental basis for the further functional investigation of miR-23a and miR-27a.
authors.affiliation: Department of Pathology, Chinese Academy of Medical Sciences, Beijing, China.
authors.forename: Yi-hui
authors.initials: YH
authors.lastname: Ma
authors.forename: Shuang-ni
authors.initials: SN
authors.lastname: Yu
authors.forename: Zhao-hui
authors.initials: ZH
authors.lastname: Lu
authors.forename: Jie
authors.initials: J
authors.lastname: Chen
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D057809:HEK293 Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D061986:MCF-7 Cells; D008565:Membrane Proteins; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics*; D014162:Transfection
chemical_list: D020413:3' Untranslated Regions; D047908:Intracellular Signaling Peptides and Proteins; C552430:MIRN23a microRNA, human; C531388:MIRN27 microRNA, human; D008565:Membrane Proteins; D035683:MicroRNAs; C485561:SPRY2 protein, human