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find Author "田洁" 1 results
  • Construction, identification and application of fusion plasmid of brainderived neurotrophic factorgreen fluorescent protein

    Objective To construct expression plasmid of the fusion protein of brainderived neurotrophic factor (BDNF)green fluorescent protein (GFP), and observe its characteristics.Methods BDNF cDNA segment was inserted into plasmid pcDNA3.1/ NT-GFP-TOPO and in the same reading frame with GFP. After verified by sequencing, the BDNFGFP plasmid was transferred into cultured Schwann cells by electroporation. And the expression of BDNFGFP fusion protein was observed by immunohistochemistry and Western blotting. The neuralprotective function of the fusion protein was evaluated by transferring the plasmid into adult rat retinas with transected optic nerve.Results The sequence of BDNFGFP plasmid was verified correctly by autosequencing. The results of Western blotting showed that the BDNF-GFP fusion protein expressed a brand with the relative molecular mass of 41times;103. Seven days after the optic nerve was transected, the number of survival retinal ganglion cells (RGC) in BDNF-GFP group and GFP group was (1201plusmn;286) and(482plusmn;151)cells/mm2, respectively; and the survival rate was (51.39plusmn;12.24)% and (20.62plusmn;6.46)% , respectively. Twentyeight days after the optic nerve was transected, the number of survival RGC in the two groups was (715plusmn;71) and (112plusmn;24)cells/mm2, respectively; the survival rate was(30.59plusmn;3.04)% and (4.79plusmn;1.03)% respectively. The differences of the survival rate of RGC between the two groups were significant (t=3.144,11.378;Plt;0.01).Conclusion BDNF-GFP fusion plasmid can express a fusion protein which emit green fluorescence and has the biological activity of BDNF.

    Release date:2016-09-02 05:43 Export PDF Favorites Scan
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