Cloning of tight junction protein claudin-1 and construction of the mammalian expression vector.
- Author:
Lin CHEN
1
;
Bo JIANG
;
Ya-li ZHANG
;
Hong-quan ZHANG
;
Wei GONG
;
Qing LU
Author Information
- Publication Type:Journal Article
- MeSH: Cell Line, Tumor; Claudin-1; Cloning, Molecular; Colorectal Neoplasms; genetics; metabolism; pathology; Genetic Vectors; genetics; Green Fluorescent Proteins; genetics; metabolism; Humans; Membrane Proteins; genetics; metabolism; Microscopy, Confocal; RNA, Messenger; biosynthesis; genetics; Recombinant Fusion Proteins; genetics; metabolism; Reverse Transcriptase Polymerase Chain Reaction; Tight Junctions; metabolism; Transfection
- From: Journal of Southern Medical University 2007;27(3):349-351
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct a recombinant plasmid containing the coding region of tight junction protein claudin-1 gene to understand the functional role of claudin-1 in human colorectal carcinoma.
METHODSThe total RNA was extracted using Trizol from human colorectal carcinoma cell line SW620, and the DNA for claudin-1 was obtained by means of RT-PCR. The PCR product was inserted into the plasmid pEGFP-C1 after restriction endonuclease digestion and ligation. The recombinant plasmid was then transfected into human colorectal carcinoma cell line SW480.
RESULTSThe sequence of the recombinant plasmid was verified by restriction endonuclease and DNA sequence analysis, and the target protein expression was detected mostly on the cell membrane.
CONCLUSIONThe expression vector claudin-1/pEGFP-C1 has been constructed successfully and the target protein can be expressed in human colorectal carcinoma cell line.
