Construction of eukaryotic expression vector of UCA1 a(CUDR) gene and its expression in bladder cancer UM-UC-2 cells / 吉林大学学报(医学版)

ABSTRACT

Objective 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-2 1 1 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-UCA1 a (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.