Abstract:

To construct an eukaryotic expression vector pcDNA-UCA1a(CUDR) and to observe its expression in bladder cancer UM-UC-2 cells,and to provide experimental basis for study on the relationship between UCA1a(CUDR) gene and bladder cancer.Methods Human total length of UCA1a(CUDR) gene was obtained from the 5′-RACE-Ready cDNA of bladder cancer BLZ-211 cells by PCR and was inserted into pcDNA3.1(＋) vector.pcDNA-UCA1a(CUDR) was identified by digestion with EcoRⅠ and BamHⅠ.The bladder cancer UM-UC-2 cells were transfected stably with the constructed eukaryotic expression vector pcDNA-UCA1a(CUDR).The expressions of UCA1a(CUDR) gene in the UM-UC-2 cells transfected with pcDNA-UCA1a(CUDR) and the UM-UC-2 cells transfected with pcDNA3.1(＋) (control vector) were detected by RT-PCR. Results The inserted fragment with 2 200 bp was successfully amplified,which was in accordance with the expected results.The eukaryotic expression vector pcDNA-UCA1a(CUDR) was constructed successfully after identified by double enzyme digestion and sequencing.The RT-PCR results showed that the expression of UCA1a(CUDR) gene in the cells transfected with pcDNA-UCA1a(CUDR) was significantly increased compared with the cells transfected with pcDNA3.1(＋).Conclusion The eukaryotic expression vector pcDNA-UCA1a(CUDR) is successfully constructed.The UCA1a(CUDR) gene highly expresses in the UM-UC-2 cells transfected with the expression vector.

Key words: UCA1a(CUDR) gene, eukaryotic expression vector, transfection, UM-UC-2 cell line