title: [Construction and screening of the artificial miRNA plasmids targeting porcine Toll-like receptor 7 gene].
abstract: OBJECTIVE
  To construct and screen specific artificial microRNA (amiRNA) expression plasmids targeting porcine Toll-like receptor 7 (TLR7) gene.
  
  
  METHODS
  The 1984-2649 bp sequence of the porcine TLR7 cDNA was amplified by RT-PCR and inserted into plasmid pEGFP-N1 to construct the fusion expression vector pTLR7-EGFP. Five amiRNAs targeting porcine TLR7 gene were designed and cloned into pcDNA5-miR to construct the recombinant interfering plasmids pcDNA5-miRTLR7. NIH-3T3 cells were co-transfected with plasmids pTLR7-EGFP and pcDNA5-miRTLR7. The expression levels of amiTLR7 were monitored by RT-PCR and their silencing efficiencies were detected by fluorescent microscopy and flow cytometry.
  
  
  RESULTS
  All five amiTLR7s were successfully constructed and could effectively silence the expression of TLR7 gene in NIH-3T3 cells with the inhibition efficiencies ranging from 36.99% to 97.28%, among which amiTLR7-3 had the best interference efficiency.
  
  
  CONCLUSIONS
  The specific artificial amiRNA expression plasmids targeting porcine TLR7 gene have been successfully constructed, and the optimal amiTLR7 with the highest inhibition efficiency has been screened.
authors.affiliation: College of Veterinary Medicine; Ministry of Education Key Laboratory of Avian Preventive Medicine, Yangzhou University, Yangzhou 225009, China. hqsong@yzu.edu.cn
authors.forename: Hongqin
authors.initials: H
authors.lastname: Song
authors.forename: Cuicui
authors.initials: C
authors.lastname: Jiang
authors.forename: Huaichang
authors.initials: H
authors.lastname: Sun
mesh_terms: D000818:Animals; D005786:Gene Expression Regulation; D051379:Mice; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D041681:NIH 3T3 Cells; D010957:Plasmids / Q000235:genetics*; D034622:RNA Interference; D013552:Swine; D051199:Toll-Like Receptor 7 / Q000235:genetics* / Q000378:metabolism
chemical_list: D035683:MicroRNAs; D051199:Toll-Like Receptor 7