Mircute rna isolation kit

miRNAs were extracted using the miRcute RNA Isolation Kit (Tiangen Biotech, Beijing, China). Quality control, labeling, and hybridization were performed commercially according to protocols in the μParaflo microRNA microarray assay (LC Sciences, Hangzhou, Zhejiang, China). Fluorescence images were collected using a laser scanner GenePix 4000B (Molecular Device, Sunnyvale, CA) and digitized using Array-Pro image analysis software (Media Cybernetics, Rockville, MD). Data were transformed by first subtracting the background and then normalizing the signals using a locally weighted regression filter.^{7 (link)}

MiRNA was extracted from microvesicles or VECs using the miRCute RNA isolation kit (TIANGEN) following the manufacturer’s instructions. The isolated RNA was reverse transcribed using the All-in-One^TM miRNA RT Detection kit (Genecopoeia, America). RT-PCR was performed using the All-in-One^TM miRNA qPCR detection kit (Genecopoeia, America) on a C1000^TM Thermal Cycler Real-Time PCR system from Applied Biosystems (Bio-Rad, United States). Fold induction was calculated using the Ct method as follows: ΔΔCt = (Ct target miRNA - Ct U6), and the final values were determined using 2^–ΔΔCt (Zhang et al., 2010 (link)).