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Title: Establishment of CHO cell line expressing human MCHR2 gene and research of its molecular<i style=""> </i>characteristics
Authors: Yuan, Chengfu
Bu, Youquan
Sengupta, Joyeeta
Yang, Junxia
Huang, Xiuning
Cheng, Li
Zhang, Qin
Yi, Faping
Liu, Geli
Song, Fangzhou
Keywords: Eukaryotic expression vector
Gene expression
Molecular characteristics
Issue Date: Nov-2009
Publisher: CSIR
Abstract: The whole length of MCHR2 gene cDNA fragment was amplified by PCR using human fetal brain cDNA library as template. The<b style=""> </b>pcDNA3.1 (+)/MCHR2 eukaryotic expression vector was constructed successfully. The recombinant pcDNA3.1 (+)/MCHR2 plasmid was transfected into Chinese hamster ovary (CHO) cell by lipofectamine<sup>TM</sup>2000, after G418 selection and then the CHO cell line expressing MCHR2 gene was established. The MCHR2 gene expression was tested by RT-PCR, western blotting and immunofluorescence. The maximum binding (B<sub>max</sub>) of CHO cell line was 309.97±1.14 f<i style="">M</i>·mg<sup>-1</sup>protein and the dissociation constant (K<sub>d </sub>value) was 0.170±0.0006 n<i style="">M</i>. MCH could stimulate Ca<sup>2+ </sup>release, its 50% effective concentration (EC<sub>50</sub>)<sub> </sub>was 2.32±0.01 n<i style="">M</i>.<b style=""> </b>The construction of the CHO cell line and the research of MCHR2 molecular characteristics have established a good experimental basis for the further research about the function of MCHR2 gene.<b style=""></b>
Description: 862-870
ISSN: 0975-1009 (Online); 0019-5189 (Print)
Appears in Collections:IJEB Vol.47(11) [November 2009]

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