November 16, 2011
Figure 1. Confocal images at 400x magnification of HeLa cells to locate GFP activity. HeLa cells were (A) tagged with Green Fluorescent Protein (GFP) (B) labeled with GFP and 14.13 µl Dexamethasone (C) tagged with GFP fused to the glucocorticoid binding domain (GBD) and (D) the subcellular localization of GFP-GBD in the presence of 14.13 µl Dexamethasone.
HeLa cells were transfected with a vector plasmid containing either GFP (A,B) or GFP-GBD (B,C). Following incubation for two days in 1 ml of complete media, the cells containing 0.45 ml of transfection complexes were washed with 1 ml HBSS to remove any background activity of the GBD domain. Figures (B) and (D) included 14.13 µl of dexamethasone whereas figure (C) was without dexamethasone. Results
A plasmid vector pRK5 has been transfected into HeLa cells. 125 ng of 210 µg/ml pRK5-GFP DNA and 100 µg/ml pRK5-GBD-GFP DNA were added to the Buffer EC in different wells at the same ratio, 0.595 µl and 1.25 µl, respectively. Two of the wells contained GFP DNA whereas the other two wells contained GFP-GBD DNA. All four of the wells were condensed with 1 µl of an enhancer and added to a lipid-based transfection reagent Effectene to allow the target DNA to be introduced into the HeLa cells. The cells containing .45 ml of transfection complexes and complete growth medium were incubated for two days and then replaced with 1 ml of HBSS and incubated for another hour. Approximately 14.13 µl of glucocorticoid dexamethasone diluted in PBS was added to one of the two wells containing GFP DNA (Figure 1 B) and a second well containing GFP-GBD DNA (Figure 1 D). Figure 1 C shows the GFB-GBD DNA lacking dexamethasone and figure 1 A displays the HeLa cells with just GFP DNA. The...
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