Confluent cells in 6-cm plates used for RT-qPCR analysis were quiesced, incubated with MRS broth or supernatants, and stimulated with Ang II according to Section 2.5 . Total RNA was extracted from cells using the FastPure®® Cell/Tissue total RNA isolation kits (Vazyme, Nanjing, China) as described by the manufacturer. The concentration was determined using a Nanodrop 2000 spectrophotometer (Thermo Fisher Inc., Waltham, MA, USA). Single-strand cDNA was synthesized from total RNA using the HiScript®® III-RT SuperMix for qPCR (+gDNA wiper) (Vazyme) according to the protocol provided. Quantitative real-time PCR (qPCR) was conducted as described in the protocol provided with iTaq™ Universal SYBR®® Green Supermix (Bio-Rad Laboratories, Inc., Shanghai, China), in a CFX ConnectTM Real-Time System (Bio-Rad Laboratories, Inc.). Real-time PCR data were analyzed using the 2−ΔΔCt method and GAPDH was used as the reference gene. Table S1 lists the qPCR primers.
Fastpure cell tissue total rna isolation kit
Manufactured by Vazyme
Sourced in China, United States
The FastPure Cell/Tissue Total RNA Isolation Kit is a product designed to efficiently extract and purify total RNA from a variety of cell and tissue samples. The kit employs a silica-based membrane technology to capture and elute the RNA, ensuring high-quality RNA suitable for downstream applications such as RT-PCR, Northern blotting, and RNA sequencing.
Automatically generated - may contain errors
Lab products found in correlation
Chamq universal sybr qpcr master mix, by Vazyme (39 mentions)
Nanodrop 2000 spectrophotometer, by Thermo Fisher Scientific (11 mentions)
Hiscript 3 rt supermix for qpcr gdna wiper, by Vazyme (11 mentions)
Hiscript 2 q rt supermix for qpcr, by Vazyme (11 mentions)
Aceq universal sybr qpcr master mix, by Vazyme (11 mentions)
Hiscript 3 rt supermix, by Vazyme (9 mentions)
Hiscript 3 rt supermix for qpcr, by Vazyme (7 mentions)
Hiscript 2 q rt supermix, by Vazyme (7 mentions)
Revertaid first strand cdna synthesis kit, by Thermo Fisher Scientific (7 mentions)
Nanodrop 2000, by Thermo Fisher Scientific (7 mentions)
Chamq sybr qpcr master mix, by Vazyme (6 mentions)
Hiscript 3 1st strand cdna synthesis kit gdna wiper, by Vazyme (6 mentions)
Hiscript 2 1st strand cdna synthesis kit, by Vazyme (6 mentions)
Quantstudio 5 real time pcr system, by Thermo Fisher Scientific (6 mentions)
Hiscript 2 q select rt supermix for qpcr, by Vazyme (5 mentions)
Primescript rt master mix, by Takara Bio (5 mentions)
Hiscript 3 first strand cdna synthesis kit, by Vazyme (4 mentions)
Aceq qpcr sybr green master mix, by Vazyme (4 mentions)
Cfx96 touch real time pcr detection system, by Bio-Rad (4 mentions)
Taq pro universal sybr qpcr master mix, by Vazyme (4 mentions)
188 protocols using fastpure cell tissue total rna isolation kit
1
Quantifying Gene Expression via RT-qPCR
2
RNA Extraction and qPCR Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
FastPure Cell/Tissue Total RNA Isolation Kits (RC101-01, Vazyme, China) were used for the total RNA extraction from mouse tissues. HiScript III RT SuperMix for qPCR (plus gDNA wiper, R323-01, Vazyme, China) was used for RNA reverse transcription, and ChamQ Universal SYBR qPCR Master Mix (Q711-02, Vazyme, China) was used for the quantitative PCR analysis of gene expression. The relative amount of the target mRNA was normalized to the RPL-19 level, and the results were calculated by the 2–ΔΔCt method. The detailed method has been described previously (Gao et al., 2017 (link)). The primer sequences are presented in Supplementary Table 2 .
3
Quantitative Analysis of Cardiac Gene Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA of hiPSC-CMs was isolated using FastPure® Cell/Tissue Total RNA Isolation Kit (Vazyme Biotech Co., Ltd, America) and their respective complementary DNA (cDNA) was then synthesized utilizing the SweScript RT I First Strand cDNA Synthesis kit (Servicebio Biotech Co., Ltd, China) as per manufacturer's instructions. Gene expression of MYH6, MYH7, MLC2a, MLC2v and cTnI were quantified from 1 μL of template cDNA using ChamQ Universal SYBR qPCR Master Mix (Vazyme Biotech Co., Ltd, America) with specific primers (Integrated DNA Technologies) and detected by LightCycler® 96 Real-time PCR system (Roche). All assays were performed in triplicate and the relative expression levels of each single gene were normalized using GAPDH and calculated using the 2 -ΔΔCt method.
4
Total RNA Extraction and qRT-PCR Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA was extracted using the FastPure Cell/Tissue Total RNA Isolation Kit (RC101-01, Vazyme, China) with on-column DNA digestion. The reverse transcription was performed using a reverse transcription kit (R333-01, Vazyme, China), with the reaction system containing 4 µL of 5 × qRT SuperMix II, 1 µg of total RNA, and RNase-free ddH2O to a final volume of 20 µL. The reaction program was set as follows: 15 min at 50 °C, 5 s at 85 °C, and then storage at 4 °C. The primers for qRT-PCR were designed using Primer Premier 5.0 and are summarized in Table S5 . GAPDH was used as control. All qPCR reactions were conducted in a 10 µL reaction volume with 1 µL of cDNA, 0.2 µL of each primer (10 µmol/L), 5 µL of 2 × AceQ Universal SYBR qPCR Master Mix, and 3.6 µL of ddH2O. Thermocycler settings were as follows: 95 °C for 5 min, 40 cycles of 95 °C for 10 s, and 60 °C for 30 s. Three replicates were conducted for all analyses.
5
Real-Time PCR Analysis of RNA Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA was extracted from various groups of 786-O cells using a FastPure Cell/Tissue Total RNA Isolation Kit (Vazyme, Nanjing, China). Complementary DNA (cDNA) was prepared from 500 ng of total RNA according to the reverse transcription protocol using the PrimeScript™ RT Reagent Kit (Takara, Dalian, China). Real-time Polymerase Chain Reaction (RT-PCR) analyses were performed using SYBR® Premix Ex Taq™ (Takara). Samples were analyzed using an ABI PRISM® 7900HT Real‐Time PCR System (Applied Biosystems, Waltham, MA, USA). The human mRNA primer sequences for target genes are listed in Table S1
6
RNA Extraction and RT-qPCR Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The total lung tissue RNA was extracted using the Fast Pure Cell/Tissue Total RNA Isolation Kit (Vazyme Biotech co., ltd., China) and converted to cDNA via reverse transcription. SYBR green on a Step One Sequence Detection System was used to perform the RT-qPCR assay. The 2−ΔΔCT formula was applied to compute the comparative abundance of genes, with β-actin as an internal control. Supplementary Table S1 detailed the primer sequences.
7
Quantitative RT-PCR for Gene Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total RNA was isolated using FastPure Cell/Tissue
Total RNA Isolation Kit (Vazyme Biotech) according to the manufacturer’s
protocol. cDNA was generated from total RNA using the PrimeScript
RT-PCR Kit (Takara). Real-time quantitative PCR was performed with
target gene-specific TaqMan probes and primers by using an Applied
Biosystems 7500 system. Each primer and probe used in this study is
listed inSupporting Information Table
S1. Relative quantification of gene expression was calculated by the
2–ΔΔCt method with
GAPDH as the endogenous reference.
Total RNA Isolation Kit (Vazyme Biotech) according to the manufacturer’s
protocol. cDNA was generated from total RNA using the PrimeScript
RT-PCR Kit (Takara). Real-time quantitative PCR was performed with
target gene-specific TaqMan probes and primers by using an Applied
Biosystems 7500 system. Each primer and probe used in this study is
listed in
S1. Relative quantification of gene expression was calculated by the
2–ΔΔCt method with
GAPDH as the endogenous reference.
8
PM2.5 Exposure Effects on Gene Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
BEAS-2B cells were treated with PM2.5 at different time points (3 hrs, 6 hrs, 12 hrs, 24 hrs and 48 hrs). The total RNA was isolated using the Fastpure Cell/Tissue Total RNA Isolation Kit (Vazyme, China) according to the manufacturer’s instructions. The purified RNA was synthesized using HiScript IIQ RT SuperMix for qPCR (Vazyme, R223–01) for the first strand cDNA. The quantitative real-time PCR analyses were performed with duplicate samples using ChamQ Universal SYBR qPCR Master Mix (Vazyme, China). The data was calculated by using 2-ΔΔCT, ACTIN as the internal reference gene.
9
Transcriptomic Analysis of Hep3B Cells Treated with D. indica
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Hep3B cells treated with 1 mg/mL D. indica and untreated Hep3B cells were used as the treatment (M1) and control groups (M0), respectively. RNA extraction was performed using the FastPure Cell/Tissue Total RNA Isolation Kit (Vazyme, RC101). Furthermore, a cDNA library was constructed using rRNA removal and a chain-specific library scheme. The Ribo-off rRNA Depletion Kit (Human/Mouse/Rat, N406) (Vazyme) was used for rRNA removal. cDNA transcription and chain-specific library construction were carried out using the VAHTS Universal V6 RNA-seq Library Prep Kit for Illumina (NR604, Vazyme). The Illumina NextSeq 500 high-throughput sequencing platform with a high-output (300 cycles) sequencing reagent was used for sequencing.
10
Quantitative Gene Expression Analysis in Lung Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The total RNA of lung tissues was extracted using a FastPure® Cell/Tissue Total RNA Isolation Kit (Vazyme Biotech Co., Ltd., Nanjing, China) following the manufacturer’s instructions. First-strand cDNA was synthesized using HiScript® III SuperMix for qPCR (Vazyme Biotech Co., Ltd., Nanjing, China), and stored at −20 °C. A real-time PCR reaction system was prepared using the iTaq Master SYBR Green Super Mix (Bio-Rad Laboratories Inc., Hercules, CA, USA) and carried out with a CFX96 Thermal Cycler (Bio-Rad Laboratories Inc.). The expressions of genes were calculated using the 2−ΔΔCT method, and normalized to that of GAPDH. Table 1 shows the primer sequences.
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!