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GPC3 and closely related to the variety of tumors, including liver cancer, malignant melanoma, ovarian cancer, breast cancer, and play different roles in different tumors, and could even be completely opposite effect. HCC, GPC3 in HCC may play an important biological function. Gene transfection techniques gene transfection to specific cells, observation of the target gene expression and cell biological characteristics, is currently the most widely used, the most mature technology, genetic research method. In this study, vector construction constructed the eukaryotic expression vector with enhanced green fluorescent protein marker GPC3 gene transfection and in vitro tumor invasion and migration assay, protein and gene level of GPC3 on the biological properties of hepatoma cells observed GPC3 two growth factors FGF2, IGF2 cell proliferation effect. One, of GPC3 green fluorescent protein true nuclear expression vectors of the building according to the GenBank Find the GPC3 open reading frame (1740bp), self-designed cited material, the application of PCR cloned by the purpose of gene GPC3 fragment PCR product with the pEGFP-N2 plasmid, respectively, by the Xho Ⅰ and EcoR Ⅰ double digestion, plastic recycling, connection, transformation and picked clones were sent to sequencing results successfully constructed with enhanced green fluorescent protein-labeled recombinant plasmid pEGFP-N2-GPC3. GPC3 expression in SK-Hep-1 hepatoma cells and of GPC3 its biological properties successfully constructed the GPC3 gene green fluorescent protein fusion expression vector pEGFP-N2-GPC3 system based on the use of liposomal transfection technology-mediated recombinant GPC3 gene expression in SK-Hep-1 cells, G418 (600μg/ml) was screened stably transfected cells, the obtained cells named GPC3-SK-Hep-1, and set the empty plasmid control group pEGFP- SK-Hep-1 and blank control SK-Hep-1, protein expression observed by fluorescence microscopy, RT-PCR detect GPC3mRNA level of expression. GPC3 gene has been successfully transfected into SK-Hep-1 cells, gene transfection group, empty plasmid control group and blank control group the morphological characteristics of the three groups of cells observed under the light environment, the results of each group no significant cell morphology differences; MTT colorimetric transgenic SK-Hep-1 cell proliferation, gene transfection group and the control group, the difference was statistically significant (P lt; 0.001) between the empty plasmid control group and blank control group , the difference was not statistically significance (P gt; 0.05) GPC3 inhibits proliferation of SK-Hep-1; adhesion assay, gene transfection group, empty plasmid control group cells and Matrigel glue adhesion rate (10.21 ± 0.62, respectively) %, (15.13 ± 0.83)%, cell adhesion rate of gene transfection group was significantly decreased (P lt; 0.001), inhibition rate was 32.52%; migration experiment 200 times eyepiece gene transfected cells transmembrane cell count was 131.7 ± 7.44. Empty plasmid control cells penetrating cell number was 71.6 ± 4.76, and the number of transmembrane cell invasion assay, 200 times the eyepiece gene transfected cells was 220 ± 12.8. Empty plasmid control group cells penetrating cell count of 138 ± 10.5. Two groups of cells relative comparison, migration and invasion abilities were significant with enhanced (P lt; 0.001); MTT assay study GPC3 growth factor FGF2 of IGF2 cell proliferation effect of GPC3 significantly with inhibition of FGF2 referral guide the cell proliferation effect (P lt ; 0.05), while the cell proliferation effect of the IGF2 mediated no change (P GT; 0.05) speculated that GPC3 in the the FGF2 signal path, play the role of the negative regulator.
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