Mirnas qpcr quantitation kit

Total RNA from tissues or transfected cells was isolated using Trizol reagent (Invitrogen) according to the manufacturer's protocol. The quality and yield of the RNA was examined by the absorbance at 260 and 280 nm. Only samples with an A260:A280 ratio between 1.8 and 2.1 were considered for further analysis. To synthesize cDNA, total RNA was reversely transcribed using prime Script RT reagent Kit (Takara, Japan), miRNAs qPCR Quantitation Kit (Genepharma, Shanghai, China), according to the manufacturer's instructions. The primers for miRNAs (miR-10a, miR-10b, miR-125b-2, miR-195, miR-21, miR-221, miR-29, miR-381, and miR-455) were purchased from RiboBio (Guangzhou, China). Quantitative real-time PCR was carried out in ABI7500 sequence detection system (Applied Biosystems, Foster City, CA, USA) using SYBR Green, according to the manufacturer's protocol. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) or U6 snRNA was used as an endogenous control. All samples were normalized to internal controls and fold changes were calculated through relative quantification (2^−ΔΔCt). The primers sequences are shown in Supplementary Table 1.

Total RNA was extracted using Trizol reagent (Invitrogen) according to the manufacturer's protocol. To quantitate miR-203 expression, total RNA was polyadenylated and reversely transcribed using miRNAs qPCR Quantitation Kit (Genepharma, Shanghai, China). To measure the mRNA levels of SNAI2, ZEB1, E-cadherin and vimentin, total RNA was reversely transcribed using primeScript RT reagent Kit (Takala, Dalian, China). Quantitative real-time PCR was carried out in ABI7500 sequence detection system (Applied Biosystems, Foster City, CA, USA) using SYBR Green according to the manufacturer's instructions. The primers were listed in Supplementary Table 3. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) or U6 snRNA was used as an endogenous control. All samples were normalized to internal controls and fold changes were calculated through relative quantification (2^−ΔΔCt).