Protocol detail

Quantitative PCR Analysis of miRNA and mRNA

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Total RNA was extracted from cells or ovarian tissue using Trizol reagent (Life Technologies). Subsequently, 2 μg of RNA was reverse-transcribed into cDNA using the PrimeScript RT reagent kit (Bio-Rad, Hercules, CA, USA), and quantitative real-time PCR (qRT-PCR) was performed on a MyiQ Single-Color Real-Time PCR Detection System (Bio-Rad). The miR-181a and mRNA expression levels were normalized to U6 small nuclear RNA and 18S, respectively, using the 2^−△△CT method as previously described.^{22 (link)} The specific primer sequences are listed in Table 1.

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Zhang M., Zhang Q., Hu Y., Xu L., Jiang Y., Zhang C., Ding L., Jiang R., Sun J., Sun H, & Yan G. (2017). miR-181a increases FoxO1 acetylation and promotes granulosa cell apoptosis via SIRT1 downregulation. Cell Death & Disease, 8(10), e3088-.

Publication 2017

Trizol reagent PrimeScript RT reagent kit MyiQ Single-Color Real-Time PCR Detection System

Cdna Mrna Ovarian Primer Quantitative real time pcr Tissue Trizol U6 small nuclear rna

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Variable analysis

independent variables

Total RNA extracted using Trizol reagent
2 μg of RNA reverse-transcribed into cDNA using PrimeScript RT reagent kit

dependent variables

MiR-181a expression levels
MRNA expression levels

control variables

U6 small nuclear RNA

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