The Construction of a Screening System of Human Amyloid Precursor Protein and Enhanced Fluorescence Protein Fusion Gene

Objective In order to screen for medicine that will act on human amyloid precursor protein(APP) at mRNA level,an enhanced fluorescence protein(EGFP) vector haboring human APP was constructed.Methods Both plasmids included pEGFP and pXCJL carring subtype APP695 digested at same a HindⅢ site to generate EGFP vector and APP695 fragment,two fragments were ligated.The possible recombinant plasmids were tested by PCR.The right orientation was identified by restriction endonuclease analysis.The recombinant plasmid was verified by sequencing.pCDNA3.1 carring APP751 was digested at XbaⅠ,blunted with Klenow fragment.Digested by HindⅢ,APP751 was recovered and ligated with pEGFP vector digested at SmaⅠand HindⅢ.The recombinant was confirmed by restriction endonuclease analysis and sequencing.COS-7 cells were transfected by recombinant plasmids to test the EGFP expression.Results EGFP carried by two recombinant plasmids could normally be expressed in COS-7 cells.Conclusions An APP gene screening system was constructed.It is quick and convenient to screen for medicine targeted APP gene at mRNA level.