Effects Of Wnt Signal Pathway On Proliferation And Differentiation Of Neural Stem Cells Derived From Adult Mouse Spinal Cord

Objective To investigate the effects of Wnt signal pathway on proliferation anddifferentiation of neural stem cells in adult mouse spinal cord.Methodsâ‘  Isolation , incubation and identification of neural stem cells from spinal cord in adult mouse.With serum free incubation (DMEM/F12+EGF+B27) and single-cell cloning technique, We isolated cells from spinal cord in adult mice. They had the capability of single-cell cloning and specific antigen of neural stem cells (Nestin) expressing was detected by Immunofluorescence technique .When employed to differentiated ,these cells expressed specific antigen of neuron(Microtubule associated protein-2;MAP-2) and astrocyts(Glial fibrillary acidic protein ;GFAP).â‘¡ Cloning, sequencing and construction of eukaryotic expression vector of mouse Wnt-3a gene.Total RNA was extracted by Trizol reagent from the pregnant 13.5d (E13.5d) Kun-Ming mouse under aseptic condition. The complete encoded sequence of mouse Wnt-3a gene had been amplified by revese transcription PCR from mouse embryo brain. Fragment was collected and purificated. Constructing pMD18-T Vector-Wnt3a clone plasmid and transformed it into competence JM109 E. coli.After preliminary screening by PCR, the positive clone was digested by double restriction enzyms MIuI and Hind III.Meanwhile, the positive clone was sequenced.Wnt-3a gene had been amplified...