MiRNA targeting gene screening technique
Cells were processed for RNA extraction, which was then isolated using Trizol (Sigma Corporation) and chloroform. The isolated RNA was precipitated with isopropanol. Any DNA present was eliminated by treating RNA with DNase I (Takara Corporation). The RNA was subsequently converted to cDNA via RT-PCR. In parallel, DH5α bacteria were cultured in 2XYT media devoid of antibiotics, followed by their amplification and harvesting. The bacterial cells were rendered competent using calcium chloride, combined with glycerol, and preserved at -80°C. Plasmid presence was confirmed through colony PCR, which encompassed preparing a PCR reaction system, programming, and initiating the PCR. Lastly, a quantitative PCR (qPCR) was executed, which involved setting up the reaction system for the first strand and primers, amalgamating and centrifuging the components, and running the qPCR.