[Expression of (His)(6)-eIF3s4 fusion protein in human breast cancer cell Bcap37].
OBJECTIVE
To construct eukaryotic expression vector of eukaryotic translation initiation factor 3 subunit 4(eIF3s4) for further functional study.


METHODS
The cDNA of eIF3s4 containing full length coding region was amplified by RT-PCR and cloned into pGEM-T Easy. The sequence of the cDNA was verified by DNA sequencing and blast against sequence data in GenBank database. The cDNA was then subcloned into the pcDNA4/HisMaxB to make a (His)(6)-eIF3s4 fusion protein expression vector. The vector was transfected into human breast cancer cell Bcap37 by Lipofectamine 2000, and the expression of (His)(6)-eIF3s4 fusion protein was detected by Western blot.


RESULTS
DNA sequencing and sequence blast showed that the cDNA amplified by RT-PCR was consistent with the eIF3s4 sequence in GenBank database, and Western blot results showed the expression of the (His)(6)-eIF3s4 fusion protein in human breast cancer cell Bcap37 as expected.


CONCLUSIONS
The (His)(6)-eIF3s4 fusion protein expression vector is constructed successfully with expression of the fusion protein in Bcap37 breast cancer cells. This work provides the basis for establishing a stable eIF3s4 expressing cell line for further study on the role of eIF3s4 in cancer multidrug resistance.
Wei
Qun
Q
Sir Run Run Shaw Hospital Clicinal Research Institute, College of Medicine, Zhejiang University, Hangzhou 310016, China.
Zhu
Feng-jia
FJ
Wang
Yi
Y
Chen
Ping
P
Wang
Lin-bo
LB
Cao
Jiang
J
D015153:Blotting, Western; D001943:Breast Neoplasms; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D039642:Eukaryotic Initiation Factors / Q000235:genetics* / Q000378:metabolism*; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D000091402:Prohibitins; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018076:DNA, Complementary; D039642:Eukaryotic Initiation Factors; C000717331:PHB2 protein, human; D000091402:Prohibitins; D011993:Recombinant Fusion Proteins