[Construction of pcDNA3.1AM and expression of adrenomedullin in mammalian cells].
The newly discovered endogenous vasodilating and diuretic peptide adrenomedullin (AM) was considered to be of attractive value in clinical treatment of hypertension and congestive heart failure. In order to explore the treatment of cardiovascular diseases by expressing AM in vivo, AM cDNA was inserted into mammalian expressing vector pcDNA3.1, and in vitro expression of AM was carried out in cultured K(562) cell line. AM mRNA was amplified by RT-PCR from the total RNA isolated from the adrenal glands of rats and was inserted into pcDNA3.1 vector to form pcDNA3.1AM, the recombinant pcDNA3.1AM was then transferred into cultured K(562) cell line by liposome. The expression of AM in pcDNA3.1AM transferred cell was identified by RT-PCR and dot immunoblot assay. The results demonstrated that there were AM mRNA in the pcDNA3.1AM-transferred K(562) cell line and AM peptides in the culturing medium, indicating that the recombinant pcDNA3.1AM vector can express AM in mammalian cell line.
Wang
Xiao-Fang
XF
Department of Physiology, Shanghai Medical College, Fudan University, Shanghai 200032.
Shao
Ying
Y
Tian
De-Zhi
DZ
Yao
Tai
T
Lu
Li-Min
LM
D053607:Adrenomedullin / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D020014:K562 Cells; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D051381:Rats; D060888:Real-Time Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection
D012333:RNA, Messenger; D011994:Recombinant Proteins; D053607:Adrenomedullin