RNA Extraction and qPCR Quantification

Freshly frozen tissues (≤30 mm³) were homogenized using the MagNA Lyser Instrument (Roche, Basel, Switzerland; 03358968001) and MagNA Lyser Green Beads (Roche; 03358941001). Total RNA from freshly frozen tissues and endometrioid adenocarcinoma cells was isolated using the RNeasy Mini Kit (Qiagen, Hilden, Germany; 74104), and complementary DNA was synthesized from genomic DNA-purified RNAs using qPCR-RT Master Mix with gDNA Remover (TOYOBO, Osaka, Japan; FSQ-301). The mRNA expression was measured using KOD SYBR qPCR Mix (TaKaRa Bio, Shiga, Japan; QKD-201X5) and QuantStudio 1 Real-Time PCR System (Thermo Fisher Scientific; A40425). Relative gene expression was analyzed using the 2^−ΔΔCt method. The primer sequences for RT-qPCR are listed in Table S2. The experiment was performed in triplicate [24 (link)].

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Kukita A., Sone K., Kaneko S., Kawakami E., Oki S., Kojima M., Wada M., Toyohara Y., Takahashi Y., Inoue F., Tanimoto S., Taguchi A., Fukuda T., Miyamoto Y., Tanikawa M., Mori-Uchino M., Tsuruga T., Iriyama T., Matsumoto Y., Nagasaka K., Wada-Hiraike O., Oda K., Hamamoto R, & Osuga Y. (2022). The Histone Methyltransferase SETD8 Regulates the Expression of Tumor Suppressor Genes via H4K20 Methylation and the p53 Signaling Pathway in Endometrial Cancer Cells. Cancers, 14(21), 5367.

Publication 2022

MagNA Lyser Instrument MagNA Lyser Green Beads RNeasy Mini Kit qPCR-RT Master Mix with gDNA Remover KOD SYBR qPCR Mix QuantStudio 1 Real-Time PCR System

Cells Complementary dna Endometrioid adenocarcinoma Frozen Gene expression Genomic Mrna Primer Rnas Tissues

Corresponding Organization : RIKEN Center for Advanced Intelligence Project

Other organizations : Chiba University, Tokyo Medical Center, National Hospital Organization, Kitano Hospital, Teikyo University

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Variable analysis

independent variables

Freshly frozen tissues (≤30 mm^3) homogenized using the MagNA Lyser Instrument (Roche, Basel, Switzerland; 03358968001) and MagNA Lyser Green Beads (Roche; 03358941001)

dependent variables

MRNA expression measured using KOD SYBR qPCR Mix (TaKaRa Bio, Shiga, Japan; QKD-201X5) and QuantStudio 1 Real-Time PCR System (Thermo Fisher Scientific; A40425)
Relative gene expression analyzed using the 2^(-ΔΔCt) method

control variables

Total RNA from freshly frozen tissues and endometrioid adenocarcinoma cells isolated using the RNeasy Mini Kit (Qiagen, Hilden, Germany; 74104)
Complementary DNA synthesized from genomic DNA-purified RNAs using qPCR-RT Master Mix with gDNA Remover (TOYOBO, Osaka, Japan; FSQ-301)

controls

No positive or negative controls explicitly mentioned

Annotations

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