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. 2004 May;15(5):2484–2491. doi: 10.1091/mbc.E03-09-0657

Figure 1.

Figure 1.

Expression of ghrelin in cells transfected with pcDNA3.1 containing ghrelin cDNA. (A) Expression of ghrelin mRNA. Total RNA was extracted from stable cell lines transfected with pcDNA3.1 or pcDNA3.1 containing ghrelin cDNAs. RT-PCR was performed using the T7 primer and the ghrelin antisense primer as indicated in the text. Lane 1, markers; lane 2, cells transfected with pcDNA3.1; lane 3, cells overexpressing ghrelin; lane 4, RT negative control; lane 5, PCR negative control; and lane 6, RT-PCR product from hypothalamus by using ghrelin sense and antisense primers. (B) Expression of ghrelin protein. Ghrelin immunoreactivity was detected by immunofluorescence in cell cultures. Control cultures lacking specific primary antibody incubation showed no significant staining. (C) Secretion of ghrelin. Five million of native 3T3 L1 cells, or cells transfected with pcDNA3.1, or cells transfected with ghrelin were cultured for 6 h. Culture media were harvested and assayed for ghrelin by using a specific rat ghrelin RIA kit. Results are expressed as mean ± SEM; n = 3. * indicates p < 0.01 as compared with native 3T3 L1 cells or cells transfected with pcDNA3.1.