Total RNA was extracted using Qiagen RNeasy mini kit from purified exosomes three days post-transduction of HEK293T cells with miR-34a virus. Revert-Aid™ First Strand cDNA Synthesis Kit (Fermentas, Germany) was applied for cDNA synthesis according to the manufacturer's instructions. The relative quantitative PCR (qPCR) experiments were performed on the ABI thermal cycler in the total volume of 20 μL containing 10 μL of Qiagen qPCR Master Mix (Qiagen), 10 pmol of each primer, and 2 μL of each cDNA. 16 s RNA was used as a reference gene.
Qpcr master mix
Manufactured by Qiagen
The QPCR master mix is a reagent used in quantitative real-time PCR (qPCR) assays. It contains all the necessary components, such as a DNA polymerase, buffers, and dNTPs, to perform qPCR amplification and detection. The master mix is designed to provide reliable and consistent results in qPCR experiments.
Automatically generated - may contain errors
Lab products found in correlation
Rneasy mini kit, by Qiagen (3 mentions)
Revertaid first strand cdna synthesis kit, by Thermo Fisher Scientific (2 mentions)
Cfx connect real time pcr detection system, by Bio-Rad (1 mentions)
Pcr array kit, by Qiagen (1 mentions)
Human tumor metastasis pcr array, by Qiagen (1 mentions)
Rt2 first strand kit, by Qiagen (1 mentions)
Trizol reagent, by Thermo Fisher Scientific (1 mentions)
Transcriptor first strand cdna synthesis kit, by Roche (1 mentions)
5 protocols using qpcr master mix
1
Quantifying miR-34a Levels in Exosomes
2
Exosomal miR-34a Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA was extracted using Qiagen RNeasy mini kit from puri ed exosomes three days posttransduction of HEK293T cells with miR-34a virus. Revert-Aid™ First Strand cDNA Synthesis Kit (Fermentas, Germany) was applied for cDNA synthesis according to the manufacturer's instructions.
The relative qPCR experiments were performed on the ABI thermal cycler in the total volume of 20 µl containing 10 µl of Qiagen qPCR Master Mix (Qiagen), 10 pmol of each primer, and 2 µl of each cDNA. 16 s RNA was used as a reference gene.
The relative qPCR experiments were performed on the ABI thermal cycler in the total volume of 20 µl containing 10 µl of Qiagen qPCR Master Mix (Qiagen), 10 pmol of each primer, and 2 µl of each cDNA. 16 s RNA was used as a reference gene.
3
mRNA Extraction and qPCR Analysis of Mitochondrial Energy Metabolism in Myocardial Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Myocardial tissue samples were homogenized in TRI Reagent (1 mL), and 200 μL chloroform was added. The sample was centrifuged at 12 000 g (4°C, 15 min), and the supernatant was transferred to a sterile centrifuge tube. Isopropanol (500 μL) was added to the supernatant, mixed, and then centrifuged again at 12 000 g (4°C, 10 min). The upper layer of transparent liquid was removed and 1 mL of 75% ethanol was added. This step was repeated 5 to 10 times. After the final ethanol wash, the sample was centrifuged at 7600 g (4°C, 5 min), placed at room temperature for 3 to 5 min, air dried, and dissolved in RNase-free water (30 μL).
mRNA was reverse transcribed to synthesize cDNA, followed by PCR. Each PCR assay used 1 μg RNA per 20 μL reaction. The experiment was performed according to the manufacturer’s instruction (PCR array kit: Mouse Mitochondrial Energy Metabolism, SABiosciences, Qiagen, PAMM-008Z). Briefly, every reaction was performed using 10 ng cDNA per 25 μL reaction volume. Quantitative PCR was performed under the following conditions: 95°C 10 min; 40 cycles of 95°C, 15 sec and 60°C, 1 min. The assays were performed on a Bio-Rad CFX ConnectTM Real-Time PCR Detection System. The qPCR master mix (QIAGEN, 330 523) was used for qPCR assay.
mRNA was reverse transcribed to synthesize cDNA, followed by PCR. Each PCR assay used 1 μg RNA per 20 μL reaction. The experiment was performed according to the manufacturer’s instruction (PCR array kit: Mouse Mitochondrial Energy Metabolism, SABiosciences, Qiagen, PAMM-008Z). Briefly, every reaction was performed using 10 ng cDNA per 25 μL reaction volume. Quantitative PCR was performed under the following conditions: 95°C 10 min; 40 cycles of 95°C, 15 sec and 60°C, 1 min. The assays were performed on a Bio-Rad CFX ConnectTM Real-Time PCR Detection System. The qPCR master mix (QIAGEN, 330 523) was used for qPCR assay.
4
Quantifying Tumor Metastasis Genes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA from HESC was purified by spin column using the RNeasy Mini Kit (Qiagen GmbH) and 0.5 µg of total RNA was used for reverse transcription (RT) with the RT2 First Strand Kit (Qiagen GmbH) according to the manufacturer's instructions. The cDNA was added to the qPCR Master Mix (Qiagen GmbH). The PCR components mix was dispensed into the Human Tumor Metastasis PCR Array format (Qiagen GmbH) according to the manufacturer's instructions. Data analysis was accomplished by the comparative Cq method (22 (link)).
5
Nrf2 RNAi in Hippocampus Transcriptome
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For Nrf2 RNAi, the mouse shRNA sequence (5′-CTTACTCTC CCAGTGAATA-3′) was subcloned into the GV248-RNAi lentiviral vector by Genechem (Shanghai, China); shRNA was expressed under the U6 promoter and green fluorescence protein (GFP) was expressed under the ubiquitin promoter. The lentiviral vectors expressing green fluorescence protein (GFP) only were used as the RNA interference (RNAi) control.
Total RNA was isolated from hippocampal tissues using TRIzol reagent (Invitrogen). mRNA was reverse-transcribed to cDNA using a Transcriptor first-strand cDNA synthesis kit (Roche) according to the manufacturer’s instructions, and real-time Q-PCR analysis was performed using the ABI StepOnePlus Real-Time PCR system (ABI VERITI96 PCR) with q-PCR Master Mix (Qiagen). Cycling conditions were as follows: 40 cycles of 95°C for 15 s, 60°C for 30 s, and 70°C for 30 s. Primer sequences are listed inTable 1 . Target gene expression was normalized to that of GAPDH. Relative gene expression was calculated with the 2–ΔΔCt formula (Figure 1 ).
Total RNA was isolated from hippocampal tissues using TRIzol reagent (Invitrogen). mRNA was reverse-transcribed to cDNA using a Transcriptor first-strand cDNA synthesis kit (Roche) according to the manufacturer’s instructions, and real-time Q-PCR analysis was performed using the ABI StepOnePlus Real-Time PCR system (ABI VERITI96 PCR) with q-PCR Master Mix (Qiagen). Cycling conditions were as follows: 40 cycles of 95°C for 15 s, 60°C for 30 s, and 70°C for 30 s. Primer sequences are listed in
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!