Total RNA from HUVECs was extracted using the RiboEX reagent (Invitrogen). One microgram of total RNA was reverse-transcribed using the Revet Aid First Strand cDNA Synthesis kit (Thermo, Rockford, IL, USA), according to the manufacturer’s instructions. Real-time PCR was performed using the SYBR Premix Ex Taq (Takara, Shiga, Japan), following the manufacturer’s protocol. All reactions were performed in triplicates. The cDNA was amplified using 60 cycles of 15 s at 95 °C, 15 s at 60 °C, and 30 s at 72 °C for each gene. The expression values are presented relative to the β-actin values in the corresponding samples. The primers used in this study are shown in Supplementary Table S1 .
Riboex reagent
Manufactured by Thermo Fisher Scientific
Sourced in United States
RiboEX reagent is a guanidinium-based solution used for the isolation and purification of total RNA from various biological samples, including cells, tissues, and bacteria. The reagent effectively disrupts cells and denatures RNases, allowing for the extraction of high-quality, intact RNA.
Automatically generated - may contain errors
Lab products found in correlation
Revet aid first strand cdna synthesis kit, by Thermo Fisher Scientific (2 mentions)
Sybr premix ex taq, by Takara Bio (2 mentions)
Nanodrop, by Thermo Fisher Scientific (2 mentions)
Reverse transcriptase, by Takara Bio (1 mentions)
Real time pcr master mix, by Biofact (1 mentions)
Stepone real time pcr system, by Thermo Fisher Scientific (1 mentions)
3 protocols using riboex reagent
1
HUVEC Total RNA Extraction and RT-qPCR
2
Cardiac Gene Expression Analysis
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Total RNA from the heart tissue was extracted using RiboEX reagent (Invitrogen). One microgram of total RNA was reverse-transcribed using the Revet Aid First Strand cDNA Synthesis kit (Thermo Fisher Scientific, Rockford, IL, USA), according to the manufacturer’s instructions. Real-time PCR was performed using SYBR Premix Ex Taq (Takara, Shiga, Japan) following the manufacturer’s protocol. All reactions were performed in triplicate. The cDNA was amplified using 60 cycles of 15 s at 95 °C, 15 s at 60 °C, and 30 s at 72 °C for each gene. The expression values are presented relative to β-actin values in the corresponding samples. The primers used in this study are listed in Supplementary Table S1 .
3
Comprehensive RNA Extraction and Analysis
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Total RNA from tissues and cultured cells was isolated using RiboEx reagent (Invitrogen, USA) following the manufacturer's protocol. The integrity and purity of the extracted total RNA were evaluated using agarose gel electrophoresis and NanoDrop (Thermo Scientific, USA). The first-strand cDNA was synthesized from total RNA (1 g) using reverse transcriptase (Takara, China). For miRNA detection, polyA tail was added to the 3 ends of RNAs before cDNA synthesis. RT-qPCR was performed using the Step One™ Real-Time PCR System (Applied Biosystems, USA) and Real-Time PCR Master Mix (Biofact, USA). The expression levels and relative expression levels were calculated using the 2 -Ct and 2 -Ct methods, respectively. The gene-specific primers are provided in Table 1. The mRNAs and miRNAs expression levels were normalized against GAPDH or U48 small RNA as the internal controls, respectively.
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