Formalin‐fixed paraffin‐embedded lung cancer tissues were obtained during surgery. Tumor specimens were procured for EGFR gene mutational analysis using previously documented methods.14 Briefly, DNA was extracted from the samples using a QIAamp DNA FFPE tissue kit (Qiagen, Hilden, Germany). EGFR mutations at exons 18–21 were analyzed, PCR amplification was performed using a Mx3000P quantitative PCR system (Stratagene; Agilent Technologies, Inc., Santa Clara, CA, USA), and data were analyzed using mxpro software version 4.10 (Stratagene; Agilent Technologies, Inc.).
Mx3000p quantitative pcr system
Manufactured by Agilent Technologies
Sourced in United States
The Mx3000P quantitative PCR system is a real-time PCR (qPCR) instrument designed for quantitative gene expression analysis. It is capable of performing multiplex PCR reactions and provides high-quality data for a wide range of applications, including gene expression profiling, pathogen detection, and microRNA analysis.
Automatically generated - may contain errors
Lab products found in correlation
Sybr premix ex taq, by Takara Bio (3 mentions)
Primescript rt reagent kit, by Takara Bio (2 mentions)
Trizol reagent, by Thermo Fisher Scientific (2 mentions)
Mxpro software version 4, by Agilent Technologies (1 mentions)
Qiaamp dna ffpe tissue kit, by Qiagen (1 mentions)
Trizol reagent, by Tiangen Biotech (1 mentions)
Sybr green realtime pcr master mix, by Toyobo (1 mentions)
Revertra ace qpcr rt kit, by Toyobo (1 mentions)
Nuclisens kit, by bioMérieux (1 mentions)
Hot firepol evagreen qpcr supermix, by Solis BioDyne (1 mentions)
Iscript cdna synthesis kit, by Bio-Rad (1 mentions)
Sybr premix ex taq, by Agilent Technologies (1 mentions)
Primescripttm rt reagent kit, by Takara Bio (1 mentions)
Rna miniprep kit, by Corning (1 mentions)
Taqman gene expression master mix, by Thermo Fisher Scientific (1 mentions)
Mxpro qpcr software, by Agilent Technologies (1 mentions)
Purelink viral rna mini kit, by Thermo Fisher Scientific (1 mentions)
Brilliant 2 qrt pcr sybr green low rox master mix, by Agilent Technologies (1 mentions)
13 protocols using mx3000p quantitative pcr system
1
EGFR Mutation Analysis in Lung Cancer
2
Quantifying EGFR Expression in Bladder Cancer
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Total RNA was extracted from four types of BCa cell sublines (5637, 5637R, T24, T24R) using TRIzol® reagent (Tiangen Biotech Co., Ltd.), according to the manufacturer's instructions. cDNA was synthesized using the ReverTra Ace™ qPCR RT Kit (cat. no. FSQ-101; Toyobo Life Science) with the following conditions: 37°C for 15 min, 98°C for 5 min and stored at 4°C. Primers for amplification of EGFR and GAPDH were designed by GenScript. Primer sequences were as follows: EGFR forward, 5′-TCCCTCAGCCACCCATATGTAC-3′ and reverse, 5′-GTCTCGGGCCATTTTGGAGAATCC-3′; GAPDH forward, 5′-CACCAACTGGGACGACAT-3′ and reverse, 5′-ACAGCCTGGATAGCAACG-3′. RT-qPCR amplification reactions were performed using BioEasy Master Mix (SYBR-Green) (cat. no. BSB30L1; Hangzhou Bioer Co., Ltd.) with a Mx3000P Quantitative PCR system (Agilent Technologies, Inc.). Amplification conditions were as follows: 95°C for 5 min, followed by 40 cycles of 95°C for 30 sec and 60°C for 45 sec. Each sample was examined in triplicate and the amount of product was normalized relative to that of GAPDH. Quantitative values were calculated according to the 2−ΔΔCq method (23 (link)).
3
Real-time PCR Analysis of GNRHR Expression
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Total RNA from transfected COS-7 cells (the GNRHR-WT, GNRHR-MUT and MOCK groups) was extracted and used for cDNA synthesis by using Rever Tra Ace® qPCR RT kit (Toyobo, Osaka, Japan). We performed Real-time PCR based on an MX3000P quantitative PCR system (Agilent, Santa Clara, CA, USA) by using SYBR® Green Realtime PCR Master Mix (Toyobo, Osaka, Japan). We performed 40 cycles of denaturing at 95 °C for 15 s, followed by annealing at 60 °C for 15 s and extension at 72 °C for 45 s. GAPDH was used as an endogenous reference. Primer sequences were as follows: GNRHR-forward: 5'-GCACGGCTGAAGACTCTAAA-3'; GNRHR-reverse: 5'-AGGCAAAGAGAAAGAAGAAGTGA-3'; GAPDH-forward: 5'-CCATCTTCCAGGAACGAGAT-3'; GAPDH-reverse: 5'-GTTGCTGACGATCTTGAGGC-3'. 2−ΔΔCt analysis was conducted to calculate the mRNA level of GNRHR.
4
Quantitative Detection of Foodborne Viruses
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On arrival, samples were thawed, shucked, and dissected. Digestive tissues (DT) were recovered, pooled, chopped, distributed in 2g aliquots, and immediately stored at −20°C until analysis. Viruses were eluted from DT using proteinase K method (ISO, 15216-1, 2017 ), and nucleic acids were extracted using the NucliSENS kit (bioMérieux, France). After validation of quality controls (extraction efficiency and inhibitor removal), the quantitative detection of norovirus and hepatitis A virus was performed by RT-qPCR (ISO, 15216-1, 2017 ). All samples were analyzed in the triplicates of undiluted nucleic acids using a one-step real-time RT-PCR kit (Invitrogen) and a MX3000P Quantitative PCR system (Agilent Technologies France).
5
Quantitative Analysis of SVCT2 Expression
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RNA from spleen/DLNs sorted-populations or in vitro cultured cells were treated for RNA isolation using TRIzol reagent (ThermoFisher, NH, USA), according to manufacture instructions. cDNA was prepared using iScript cDNA synthesis kit (Bio-Rad, CA, USA) and the expression of SVCT2 and housekeeping gene 18S was measured by real-time PCR, using HOT FIREPol EvaGreen qPCR Supermix (Solis BioDyne, Tartu, Estonia) with Mx3000P quantitative PCR system (Agilent Technologies, CA, USA). Primer sequences are showed in Table 1 .
6
Quantitative PCR Analysis of Inflammatory Genes
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Following treatment, HUVECs (1×105 cells/well) were washed twice with ice-cold PBS, and total RNA was extracted using TRIzol® reagent according to the manufacturer's instructions. The concentration of the total RNA was determined by measuring the absorbance at 260 nm. Subsequently, total RNA was reverse-transcribed into cDNA using a PrimeScript™ 1st strand cDNA Synthesis Kit. The cDNA of the target genes was amplified using the SYBR® Premix Ex Taq on the Mx3000P quantitative PCR system (Stratagene; Agilent Technologies, Inc.). The primer sequences used were as follows: Human IL-1β forward, 5′-CAT TGA GCC TCA TGC TCT GTT-3′ and reverse, 5′-CGC TGT CTG AGC GGA TGA A-3′; human IL-6 forward, 5′-TTC GGT CCA GTT GCC TTC TC-3′ and reverse, 5′-TCA CCA GGC AAG TCT CCT CA-3′; human TNF-α forward, 5′-GCT GCA CTT TGG AGT GAT CG-3′ and reverse, 5′-GCT TGA GGG TTT GCT ACA ACA-3′; and human GAPDH forward, 5′-TGT GGG CAT CAA TGG ATT TGG-3′ and reverse, 5′-ACA CCA TGT ATT CCG GGT CAA T-3′. The expression levels of the target mRNAs were normalized to those of GAPDH. All samples were run in triplicate and analyzed using the 2−ΔΔCq method as previously described (20 (link)).
7
Adenovirus Infection in Cell Cultures
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Real-time PCR was performed in cultured cells on the 1st, 3rd, 5th, 7th, 10th and 14th day after infection with adenovirus. RNA was extracted with a multisource RNA miniprep kit (Corning, NY, USA). Total RNA (500 ng) was reversely transcribed into cDNA using the PrimeScript™ RT reagent kit (TaKaRa, Dalian, China). Real-time PCR for each sample was performed on an MX3000P quantitative PCR system (Agilent, Santa Clara, CA, USA) using SYBR Premix Ex Taq (TaKaRa, Dalian, China) and the following primers: iNOS_ 5′-AAGCACATTTGGCAATGGAGAC-3′, β-actin_ 5′-GACGGTGTGCACCAACATCTA-3′, 5′-TTCTTGGCTTTCAGGATGGAG-3′. PCR condition included 95 °C for 30 s, followed by 40 cycles of 95 °C for 5 s, 60 °C for 30 s and 72 °C for 30 s. Relative expression level of target genes was calculated with 2−ΔΔCt method. β-actin was chosen as normalization control.
8
Quantitative Analysis of Telomerase Expression
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Total RNA was extracted using a multisource RNA miniprep kit (Corning, NY, USA) converted to cDNA using the PrimeScriptTM RT reagent kit (TaKaRa, Dalian, China). Real-time quantitative polymerase chain reaction (PCR) was performed using SYBR Premix Ex Taq (TaKaRa) on an MX3000P quantitative PCR system (Agilent, Santa Clara, USA). The following cycling conditions were used: pre-denaturation at 95°C for 30 seconds; 40 cycles at 95°C for 5 seconds, 60°C for 30 seconds and 72°C for 30 seconds. The level of β-actin mRNA expression was measured as an endogenous reference and used for normalization purposes. Relative quantification of the expression levels of each transcript for each group was calculated using the 2−ΔΔCt method. We used the following PCR primers: hTERT_s: 5′-GACGGTGTGCACCAACACATCTA-3, hTERT_a: 5′-TTCTTGGCTTTCAGGATGGAG-3′, rat TERT (rTERT)_s: 5′-GCTGGACACTCGGACTTTGGA-3′, rTERT_a: 5′-ACTTCAACCGCAAGACTGACAAGA-3′, β-actin_s: 5′-GACGGTGTGCACCAACATCTA-3′, β-actin_a: 5′-TTCTTGGCTTTCAGGATGGAG-3′.
9
Quantification of Oxidative Stress Genes
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Real-time PCR was performed on a MX3000P Quantitative PCR System (Agilent Technologies). Taqman primer and probe sets for genes nNos, GPX1, and GAPDH were purchased (Applied Biosystems), and PCR was performed with Taqman Gene expression master mix for primer and probe sets, according to manufacture protocol. Gapdh was used for normalization of expression of nNos and GPx-11. Fold changes were calculated in the Agilent Technologies software program, which compared experimental groups to Sham controls.
10
Quantitative Analysis of Gene Expression
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Reverse transcriptase reactions were performed with reagents from the Taqman Reverse Transcriptase Reagents Kit (Applied Biosystem) following manufacturer’s protocols. Real time PCR was performed on a MX3000P Quantitative PCR System (Agilent Technologies). Taqman primer and probe sets for genes nNos, GPx-1, and Gapdh were purchased from (Applied Biosystems), and PCR was performed with Taqman Gene expression master mix, primer sets, and probe sets according to manufacturer’s protocols. Gapdh was used for normalization of expression of nNos and GPx-1. Fold changes were calculated in the Agilent Technologies software program, which compared each transfection group to a negative control.
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