[Construction of cyclin D1 recombinant plasmids and its expression in human nasopharyngeal carcinoma cells].
OBJECTIVE
To construct the eukaryotic expression vectors of human cyclin D1 gene and express them in poorly differentiated nasopharyngeal carcinoma cells (CNE-2Z cells).


METHODS
The full-length cyclin D1 was cloned from CNE-2Z cells by RT-PCR. The cDNA fragments were inserted into pIRES2-EGFP plasmids and pEGFP-C2 plasmids and confirmed by restriction enzyme digestion, PCR and sequencing. The recombinant vectors were transfected into CNE-2Z cells via Lipofectamine 2000, and the expression of cyclin D1 in the cells was examined by immunofluorescence and Western blotting.


RESULTS
Agarose gel electrophoresis showed a 918 bp band of the RT-PCR products, which matched the expected size. Restriction enzyme digestion, PCR and sequencing demonstrated successful construction of the recombinant vectors. CNE-2Z cells transfected with the recombinant vectors expressed cyclin D1 protein or cyclin D1-GFP protein as were verified by immunofluorescence and Western blotting.


CONCLUSIONS
We have cloned cyclin D1 gene and constructed its eukaryotic expression vectors that can be expressed in nasopharyngeal carcinoma cells, which may facilitate the study of the role of cyclin D1 in the development of nasopharyngeal carcinoma.
Department of Physiology, Medical College of Jinan University, Guangzhou 510632, China.
Zhi-quan
ZQ
Bai
Hai-feng
HF
Zhang
Li-xin
LX
Chen
Lin-yan
LY
Zhu
Wan-hong
WH
Zuo
Shu-tong
ST
He
Li-wei
LW
Wang
D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D019938:Cyclin D1 / Q000096:biosynthesis* / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D009303:Nasopharyngeal Neoplasms / Q000378:metabolism / Q000473:pathology*; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection
D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D019938:Cyclin D1; D049452:Green Fluorescent Proteins