Construction of a hepatocellular carcinoma cell line that stably expresses stathmin with a Ser25 phosphorylation site mutation.
We constructed hepatocellular carcinoma (HCC) cells that stably express stathmin with a Ser25 phosphorylation site mutation (stathmin S25A). We used the polymerase chain reaction for site-directed mutagenesis, constructed a stathmin S25A plasmid, and verified the results by restriction enzyme cleavage and sequencing technology. Using the liposome transfection method, stathmin wild-type and S25A HCCLM6 cells were established, which were identified by western blotting. The sequencing report of the stathmin S25A plasmid showed that stathmin serine at position 25 had mutated into alanine. Stable cells transfected with stathmin wild-type and S25A plasmids were constructed. Using western blotting, we confirmed that the expression level of stathmin pS25 in the stathmin S25A cells was reduced than that in the stathmin wild-type and HCCLM6 control cells (P < 0.05). We constructed stathmin S25A HCCLM6 cells, which offer an experimental model for further investigation of the molecular mechanism of stathmin phosphorylation in hepatocarcinogenesis.
Department of Chemistry, Baisc Medical College, Sichuan Medical University, Luzhou, Sichuan, China.
J
J
Du
Research Center for Preclinical Medicine, Sichuan Medical University, Luzhou, Sichuan, China.
Z H
ZH
Tao
Research Center for Preclinical Medicine, Sichuan Medical University, Luzhou, Sichuan, China.
J
J
Li
Liver Cancer Research Institute, Zhongshan Hospital, Fudan University, Shanghai, China.
Y K
YK
Liu
Research Center for Preclinical Medicine, Sichuan Medical University, Luzhou, Sichuan, China.
L
L
Gan
D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism*; D050777:Stathmin / Q000235:genetics / Q000378:metabolism*
D050777:Stathmin; D012694:Serine