Construction of eukaryotic plasmid of human β-catenin gene and the expression and localization of its fusion protein

Objective To construct the expression plasmid of beta-catenin(or β-catenin) gene and identify the expression and localization of its fusion protein. Methods Total mRNA was extracted from NIH3T3 cells, cDNA was formed by reverse transcription. The β-catenin coding sequence was amplified by polymerase chain reaction(PCR) and subcloned into pEGFP-C1 vector. After the target region was identified by enzyme digestion and sequencing, the plasmid was transfected into NIH3T3 cells. The expression of the recombinant plasmid in NIH3T3 cells was detected by Western blot. The localization of pEGFP-β-catenin in NIH3T3 cells was observed with laser scanning confocal microscopy. Results The length of the fragment identified by restriction enzyme digestion was 2346 bp. The expression of pEGFP-β-catenin fusion protein with a molecular weight of 115 kDa was detected by Western blot. The pEGFP-β-catenin fusion protein was mostly localized in the membrane and cytoplasm of NIH3T3 cells. Conclusion The recombinant plasmid of β-catenin gene was successfully cloned into eukaryotic expressing vector, and the pEGFP-β-catenin fusion protein was mostly localized in the membrane and cytoplasm of NIH3T3 cells.