Construction and identification of plasmid mediated RNA interference targeting Nogo-66 receptor
- VernacularTitle:Nogo-66受体RNA干扰质粒的构建及其干扰效率鉴定
- Author:
Youming FAN
;
Mingchun CAI
- Publication Type:Journal Article
- Keywords:
Nogo-66 receptor;
RNA interference;
plasmid
- From:Journal of Third Military Medical University
2003;0(13):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct and identify of a plasmid mediated high efficiency of RNA interference(RNAi) against Nogo-66 receptor(NgR).Methods After cloning of NgR by RT-PCR,the fragments were inserted into pcDNA3.1/CT-GFP-TOPO to produce plasmid expressing NgR-GFP fusion protein.Four pairs of oligonucleotide were designed according to NgR sequence and annealed.The resulting fragments were ligated into short hairpin RNA(shRNA) expressing plasmid.The plasmids expressing NgR-GFP fusion protein and shRNA were co-transfected into AAV-293 cells.RNAi efficiency against NgR was observed under fluorescent microscope and calculated by Western blotting.Results The interference efficiency of one sequence was above 90%.Conclusion A plasmid mediated high RNAi efficiency against NgR is constructed successfully.
