Part:BBa_K3730013:Experience

The data we obtained were processed the results of which are shown in Figure. 1.

                 
               
                   Figure. 1 Fluorescence intensity of cells after transfection with 3u and 3ue.
(3u refers to 3’UTR-pcDNA3.1-N-eGFP-C, 3u refers to 3’UTRempty-pcDNA3.1-N-eGFP-C. )
               

The results showed that in Hek293T transfected with p3u, the expression of eGFP was inhibited, which was not significant enough in HepG2. (p<0.1) It demonstrated that the miRNA concentration in normal cells were likely to be sufficient to inhibit the E1A.

We also designed plasmids called pRNAiU6.2-22, pRNAiU6.2-195, pRNAiU6.2-199a to increase the quantity of these miRNA in HepG2 as described previously. That was how we could know if the effect of 3UTR was actually caused by miRNA or other variables. Thus, we transfected 3u and one of these plasmids above into HepG2. The group called 3u+u6 was the controlled group, and u6 referred to pRNAiU6 without extra part. The results are shown in Figure. 3-5. Comparatively, the expressions of eGFP were all significantly decreased in HepG2 when these miRNA were replenished in cells, mir195 was the most effective one of these three. It showed that the function of 3u was related to the quantity of these miRNA, and it was reasonable to control the expression of E1A through the controlling of miRNA content.

                 
    
                   Figure. 2 Fluorescence of cells after transfection of 3u and mir199a. mir195, mir22 or u6. 
HepG2 transfected with 3u and certain miRNAs had significantly lower expression of eGFP compared with the control group.
           
               

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