title: [Construction of miRNA expression vector dual-targeting on HIF-1α/survivin genes and its effects on proliferation of pancreatic cancer Panc-1 cell line].
abstract: To design and construct miRNA expression vector dual-targeting on HIF-1α and survivin genes and to investigate its effects on proliferation of human pancreatic cancer cells.
  
  
  
  The specific pre-miRNA single strand DNA oligos for HIF-1 α and survivin genes were designed and synthesized, then via annealing and ligating with pcDNA6.2-GW/EmGFP-miR plasmids in order, two kinds (eight in total) of miRNA expression vectors for HIF-1α and survivin genes were constructed. The vectors, which were most effective to knockdown target genes, were screened with real-time RT-PCR and combined by chaining technology to construct dual-targeting plasmid. The recombined dual-targeting plasmid, mono-targeting plasmids and negative plasmid were transfected into Panc-1 cells, the suppression effect on two genes was identified by real-time RT-PCR, Western blot and MTT assays.
  
  
  
  The miRNA expression plasmids anti-H, anti-S and anti-H+S were successfully constructed by identification of sequencing analysis, and they were able to effectively inhibit the target genes expressing. MTT assays showed that the inhibition effect of dual-targeting vector anti-H+S was higher than that of mono-targeting vectors anti-H and anti-S 72 h after transfection (P<0.05).
  
  
  
  The effective miRNA expression vector dual-targeting on HIF-1α and survivin genes has been successfully constructed. The inhibition effect on proliferation of pancreatic cancer Panc-1 cells by dual-targeting plasmid was higher than that by mono-target plasmids.
authors.affiliation: Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310016, China.
authors.forename: Sun-bing
authors.initials: SB
authors.lastname: Xu
authors.forename: Yi-ping
authors.initials: YP
authors.lastname: Zhu
authors.forename: Yi-ping
authors.initials: YP
authors.lastname: Mou
authors.forename: Ling-hua
authors.initials: LH
authors.lastname: Zhu
mesh_terms: D045744:Cell Line, Tumor; D049109:Cell Proliferation; D005822:Genetic Vectors; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics* / Q000378:metabolism; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics* / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics*; D010190:Pancreatic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*; D010957:Plasmids / Q000235:genetics; D000077022:Survivin; D014162:Transfection
chemical_list: C107489:BIRC5 protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D051034:Inhibitor of Apoptosis Proteins; D035683:MicroRNAs; D000077022:Survivin