Qsybr green containing pcr kit

Total RNA was extracted with the TRIzol reagent (Invitrogen, Carlsbad, CA, USA). LDHA cDNA was synthesized from 1 μg of total RNA using the Reverse Transcription System (Promega, Madison, WI, USA). β-actin was amplified in parallel as an internal control. For miR-200c, reverse transcription and qRT-PCR were performed using a qSYBR-green-containing PCR kit (GenePharma, Shanghai, China). U6 snRNA was used as an endogenous control. Gene expression was quantified by measuring cycle threshold (Ct) values and normalized to β-actin or U6 snRNA using the 2^−ΔΔCt method.

Total RNAs were extracted with TRIzol reagent (Invitrogen). For the detection of LDHA mRNA, cDNA was synthesized from 1 ug of total RNA using the reverse reaction kit according to the manufacturer's instructions (Promega). β-actin was amplified in parallel as an internal control. The primers were: β-actin forward: 5′-AGCGAGCATCCCCCAAAGTT-3′ and reverse: 5′-GGGCACGAAGGCTCATCATT-3′. LDHA forward: 5′-TTGGTCCAGCGTAACGTGAAC-3′ and reverse: 5′-CCAGGATGTGTAGCCTTTGAG-3′. For miR-200b, reverse transcription and qRT-PCR reactions were performed using a qSYBR-green-containing PCR kit (GenePharma, Shanghai, China). U6 snRNA was used as an endogenous control for miRNA detection. The expression of each gene was quantified by measuring cycle threshold (Ct) values and normalized using the 2^−ΔΔCt method relative to U6 snRNA or β-actin.

Total RNA was extracted from cells using TRIzol reagent (Invitrogen, USA). For miR-124, reverse transcription and qRT-PCR reactions were performed using a qSYBR-green-containing PCR kit (GenePharma, Shanghai, China). U6 snRNA was used as an endogenous control. The precursor form of miR-124 was amplified. For detecting PRRX1 mRNA, cDNA was synthesized from 1 μg of total RNA using the reverse transcription reaction kit according to the manufacturer's instructions (Promega, USA). Human GAPDH was amplified in parallel as an internal control. The primers were listed in Supplementary Table S1. All these samples were normalized to internal controls and fold changes were calculated through relative quantification (2-ΔΔCt).