Part:BBa_K4789004

1.miR-22-sponge-pepper description We designed this part with pepper fluorescence to monitor the expression of miR-22 in cells. The “Pepper” plasmid containing the part sequence of LncRNA MALAT1 in order to reflect miRNA expression in vivo. lncRNAs can interact with miRNAs as “sponges”. And the fluorescence reflect the miRNA expression in the cervical cancer in turn. We tested the sensitivity and specificity of the vectors in Hela cells. In the future, the miRNA-LncRNA MALAT1 complex could be used to screen and detect the cervical cancer. The patients could benefit from our work. The model of the plasmid was listed below (Fig 1).

                   
                        Fig 1. The diagram of miR-22- sponge-pepper

2.1 Function verification of miR-22 sensor In order to test the ability of miR22 sensor, we transfected miR-22-sponge-pepper and pre-miR-22 (overexpress miR-22 in cells) into Hela cells. The control group only transfected with miR-22-sponge-pepper (2 ug), the experimental group transfected with both miR-22-sponge-pepper (2 ug) and pre-miR-22 (1 ug). 48 hours later, we add 2 μm HBC fluorescent dye into per well. After incubation for 2 hours, cells were harvested and the green fluorescence was measured by plate reader (SpectraMax i3). The result showed that miR-22 could inhibit the fluorescence of Pepper in cells transfected with miR-22-sponge-pepper (Fig 2). The result suggested that miR-22 sensor can detect the alteration of miR-22 expression in cells.

Fig 2. The images of Hela cells transfected with different plasmids. (A). miR-22-sponge-pepper were transfected into Hela cells. (B) miR-22-sponge-pepper and pre-miR-22 were transfected into Hela cells