Validating miR-27 and SPRY2 mRNA Binding

Luciferase reporter assays were used to validate the direct binding of miR-27 and SPRY2 mRNA 3′UTR fragments as previously described [26 (link)]. In brief, wild-type and mutant SPRY2 mRNA 3′UTR fragments were cloned into the pGL3-basic dual luciferase reporter vectors (Promega, Madison, WI, U.S.A.), respectively. A dual-luciferase reporter system (BD Bioscience) was used to determine the relative luciferase activity at 48 h post-transfection. Firefly luciferase activity was normalized to Renilla luciferase activity.

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Che F., Wan C., Dai J, & Chen J. (2019). Increased expression of miR-27 predicts poor prognosis and promotes tumorigenesis in human multiple myeloma. Bioscience Reports, 39(4), BSR20182502.

Publication 2019

pGL3-basic dual luciferase reporter vectors dual-luciferase reporter system

Assays Firefly luciferase Luciferase Mrna Pgl3 Promega Renilla luciferase Transfection Vectors

Corresponding Organization : Sichuan Academy of Medical Sciences & Sichuan Provincial People's Hospital

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Variable analysis

independent variables

Wild-type SPRY2 mRNA 3′UTR fragments
Mutant SPRY2 mRNA 3′UTR fragments

dependent variables

Relative luciferase activity at 48 h post-transfection

control variables

Firefly luciferase activity was normalized to Renilla luciferase activity

controls

Positive control: wild-type SPRY2 mRNA 3′UTR fragments
Negative control: mutant SPRY2 mRNA 3′UTR fragments

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