Construction of eukaryotic expression vector for mouse PS1-GFP
Mutations in presenilin 1(PS1) gene are genetically linked to the early onset of familial Alzhemer’s disease (EOFAD). Full-length PS1 is a multiple tansmembrane protein which relates toγ-secretase and underges endoproteolytic
processing of lots of proteins , is involved in Wnt/wingless signaling, cell adhesion, cell-cell interaction and cell fate determinaton during development. But the exact structure and functions of PS1 is not known. The expression vector for mouse PS1-GFP fusion protein was constructed for providing the possibility of further investigation. Method: We got the full-length cDNA of PS1 using RT-PCR from 9d embryo and cloned the PS1 gene to pMD 18-T Vector. Then we construct the fusion protein GFP-PS1 (recombinant plasmid pEGFP-C1-PS1) by PCR using the primer (5'-GCC GAA TTC TAT GAC AGA GAT ACC TGC ACC-3', 5'-GAA GGA TCC GAT ATA AAA CTG ATG GAA TGC-3'). The sequence of primer contains EcoRI and BamHI sites. The integrity of all PCR products was confirmed by DNA sequencing. PS1-GFP fusion protein (recombinant plasmid pEGFP-N2-PS1) was constructured by enzymolysis of pMD 18-T-PS1 with EcoRI and Sa1I .