Quantifying Transcriptomic Profiles using RT-qPCR

We applied the miRcute Plus miRNA First-Strand cDNA Synthesis Kit (Tiangen, KR21) to reverse the total RNA and the miRcute Plus miRNA qPCR Detection Kit (SYBR Green; Tiangen, FP411) reagent to detect miRNA expression levels. We used HiScript II Q Select RT SuperMix for qPCR (+gDNA wiper; Vazyme, R223) to complete RNA reversal and used AceQ qPCR SYBR Green Master Mix (Vazyme, Q111) to quantify mRNA, lncRNA, and circRNA expression. The relative expression of genes was analyzed by the 2^−△△Ct method. U6 was an internal control of miRNAs (Fu et al., 2021 (link); Woo et al., 2022 (link)). β-actin was internal control of mRNAs, lncRNAs, and circRNAs (Primer sequence listing is shown in Supplementary Table S7).

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Dong W., Wang G., Bai Y., Li Y., Huo X., Zhao J., Lu W., Lu H., Wang C., Wang X., Chen H, & Tan C. (2023). Analysis of the noncoding RNA regulatory networks of H37Rv- and H37Rv△1759c-infected macrophages. Frontiers in Microbiology, 14, 1106643.

Publication 2023

miRcute Plus miRNA First-Strand cDNA Synthesis Kit miRcute Plus miRNA qPCR Detection Kit (SYBR Green; HiScript II Q Select RT SuperMix for qPCR AceQ qPCR SYBR Green Master Mix

Actin Cdna Circrnas Expression genes Lncrnas Mirnas Mrnas Primer Sybr green Synthesis

Corresponding Organization :

Other organizations : Huazhong Agricultural University, China Animal Disease Control Center

Top 5 similar protocols

Variable analysis

independent variables

MiRcute Plus miRNA First-Strand cDNA Synthesis Kit (Tiangen, KR21)
MiRcute Plus miRNA qPCR Detection Kit (SYBR Green; Tiangen, FP411)
HiScript II Q Select RT SuperMix for qPCR (+gDNA wiper; Vazyme, R223)
AceQ qPCR SYBR Green Master Mix (Vazyme, Q111)

dependent variables

MiRNA expression levels
MRNA expression
LncRNA expression
CircRNA expression

control variables

U6 as internal control for miRNAs
β-actin as internal control for mRNAs, lncRNAs, and circRNAs

positive controls

Not specified

negative controls

Not specified

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