[Cloning of human Nanog gene and its expression in COS-7L cells].
OBJECTIVE
To clone human Nanog gene and express it in mammalian cells.


METHODS
LA-PCR was used to amplify human Nanog gene from genomic DNA of HEK 293 cells. The Nanog gene was inserted into the Flag-tagged pCMV vector by gene recombination technique. After being confirmed by sequencing, the pFlag-Nanog was transfected into COS-7L cells. Western blot and indirect immunofluorescent staining against the Flag tag were used to detect the expression of Nanog protein.


RESULTS
The sequence of the cloned Nanog was confirmed to be correct by sequence analysis. The COS-7L cells transfected with pFlag-Nanog could efficiently express Nanog protein.


CONCLUSIONS
The cloning of human Nanog gene and the construction of its eukaryotic expression vector were successful. The study will lay the foundation for further research on the function of Nanog and its role in neural stem cells.
Liu
Xia
X
Institute of Neuroscience, Fourth Military Medical University, Xi'an 710032, China. Liu123xia5798@sohu.com
Long
Mei
M
Jiang
Hui
H
Jin
Wei-lin
WL
Jiao
Xi-ying
XY
Ju
Gong
G
D000818:Animals; D019556:COS Cells / Q000378:metabolism; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics*; D018398:Homeodomain Proteins / Q000096:biosynthesis* / Q000235:genetics*; D006801:Humans; D007668:Kidney / Q000166:cytology; D000071317:Nanog Homeobox Protein; D017421:Sequence Analysis
D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; C486780:NANOG protein, human; D000071317:Nanog Homeobox Protein; D004247:DNA