Sensifast syber

Manufactured by Meridian Bioscience

Sourced in Italy, United Kingdom

SensiFast Syber is a real-time PCR (polymerase chain reaction) reagent kit developed by Meridian Bioscience. It is designed for the detection and quantification of DNA and RNA sequences.

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Lab products found in correlation

Tri reagent, by Merck Group (4 mentions) Sensifast cdna synthesis kit, by Meridian Bioscience (4 mentions) Icycler iq real time detection system, by Bio-Rad (3 mentions) Cfx opus 96 real time pcr instrument, by Bio-Rad (1 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

SensiFAST SYBER® No-ROX kit (5 citations) SensiFAST SYBER Lo-ROX mix (2 citations) SensiFAST SYBER HI-ROX Kit (2 citations)

5 protocols using sensifast syber

RNA Extraction and Real-Time PCR Analysis

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Total RNA extraction was performed by using the TRI Reagent (Sigma-Aldrich, product code T9424) following the manufacturer’s instruction. For first-strand synthesis of cDNA, 1 μg of RNA was used in a 20-μl reaction mixture by using a SensiFast cDNA synthesis kit (Bioline). For real-time PCR analysis, 0.4 μl of cDNA sample was amplified by using the SensiFast Syber (Bioline) in an iCycler iQ Real-Time Detection System (Bio-Rad Laboratories GmbH, Segrate, Italy).
The reaction conditions were 95°C for 2 min followed by 40 cycles of 5 s at 95°C and 30 s at 60°C. Actin was used as internal control. The full list of primers used is reported in Supplementary Materials and Methods.

Avolio R., Järvelin A.I., Mohammed S., Agliarulo I., Condelli V., Zoppoli P., Calice G., Sarnataro D., Bechara E., Tartaglia G.G., Landriscina M., Castello A., Esposito F, & Matassa D.S. (2018). Protein Syndesmos is a novel RNA-binding protein that regulates primary cilia formation. Nucleic Acids Research, 46(22), 12067-12086.

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Total RNA Extraction and Real-Time PCR Analysis

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Total RNA extraction procedures were performed by using TRI Reagent (Merck Life Science S.r.l., Milano, Italy; product code T9424), following the manufacturer’s instruction. For first-strand synthesis of cDNA, 1 μg of RNA was used in a 20-μL reaction mixture by using a SensiFast cDNA synthesis kit (Bioline, London, UK). For real-time PCR analysis, 0.4 μL of cDNA sample was amplified by using the SensiFast Syber (Bioline, London, UK) in an iCycler iQ Real-Time Detection System (Bio-Rad Laboratories GmbH, Segrate, Italy). The reaction conditions were 95 °C for 2 min followed by 40 cycles of 5 s at 95 °C and 30 s at 60 °C. Actin was chosen as the internal control. In PCR analyses performed upon TRAP1 silencing, RNAs were collected 72 h after siRNA transfection. The following primers (Table 1) were used for PCR analysis.

Criscuolo D., Avolio R., Calice G., Laezza C., Paladino S., Navarra G., Maddalena F., Crispo F., Pagano C., Bifulco M., Landriscina M., Matassa D.S, & Esposito F. (2020). Cholesterol Homeostasis Modulates Platinum Sensitivity in Human Ovarian Cancer. Cells, 9(4), 828.

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Quantifying Gene Expression by RT-qPCR

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Total RNA extraction was performed using the TRI Reagent (Merck, product code T9424) following the manufacturer’s instruction. For first-strand synthesis of cDNA, 1μg of RNA was used in a 20-μl reaction mixture by using a SensiFast cDNA synthesis kit (Bioline). For real-time PCR analysis, 0.4μL of cDNA sample was amplified by using the SensiFast Syber (Bioline) in an CFX Opus 96 Real-Time PCR Instrument (Bio-Rad Laboratories GmbH, Segrate, Italy). The reaction conditions were 95 °C for 5 min followed by 45 cycles of 15 sec at 95 °C and 1 min at 60 °C. The sequence of the oligos used are listed below in 5’-3’ direction:

Avolio R., Agliarulo I., Criscuolo D., Sarnataro D., Auriemma M., Pennacchio S., Calice G., Ng M.Y., Giorgi C., Pinton P., Cooperman B., Landriscina M., Esposito F, & Matassa D.S. (2023). Cytosolic and mitochondrial translation elongation are coordinated through the molecular chaperone TRAP1 for the synthesis and import of mitochondrial proteins. bioRxiv.

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Investigating vATPase Role in Phagosome Maturation

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To investigate the role of vATPase in phagosome maturation, sub-confluent A549 lung epithelial cells were transfected with 500 pg of siRNA ATP6V0A2 (Ambion, Austin, TX, USA) using Lipofectamine 3000 (Invitrogen, Darmstadt, Germany) according to the manufacturer’s instructions. Cells at 48 h post-transfection were used for infection experiments and changes in the expression of vATPAse were evaluated by qPCR using SensiFAST Syber (Bioline, Memphis, TN, USA) according to the manufacturer’s instructions. Validated primers targeting ATP6V0A2 and RPL13A were purchased from Bio-rad (Hercules, CA, USA).

Ben-Ghazzi N., Moreno-Velásquez S., Seidel C., Thomson D., Denning D.W., Read N.D., Bowyer P, & Gago S. (2021). Characterisation of Aspergillus fumigatus Endocytic Trafficking within Airway Epithelial Cells Using High-Resolution Automated Quantitative Confocal Microscopy. Journal of Fungi, 7(6), 454.

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RNA Extraction and Real-Time PCR Analysis

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Total RNA extraction procedures were performed by using TRI Reagent (Merck Life Science S.r.l., Milano, Italy; product code T9424), following the manufacturer’s instruction. For first-strand synthesis of cDNA, 1 μg of RNA was used in a 20-μL reaction mixture by using a SensiFast cDNA synthesis kit (Bioline, London, UK). For real-time PCR analysis, 0.4 μL of cDNA sample was amplified by using the SensiFast Syber (Bioline, London, UK) in an iCycler iQ Real-Time Detection System (Bio-Rad Laboratories GmbH, Segrate, Italy). The reaction conditions were 95 °C for 2 min followed by 40 cycles of 5 s at 95 °C and 30 s at 60 °C. PPIA was chosen as the internal control. The following primers were used for PCR analysis:

PPIA:	Forward: 5′-CTGCACTGCCAAGACTGA-3′;	Reverse: 5′-GCCATTCCTGGACCCAAA-3′
CYC1:	Forward: 5′-TACGGACACCTCAGGCAGTG-3′;	Reverse: 5′-CACGGTGAGACCACGGATAG-3′
TTC19:	Forward: 5′-TTTGCATGACGCTCTTCGTC-3′;	Reverse: 5′-TGCATTGTCCTCCTGCTTCAT-3′
UQCRC2:	Forward: 5′-CTTACCGGAATGCCTTGGCT-3′;	Reverse: 5′-GATAAACCAAGCCCACCCCT-3′

Matassa D.S., Criscuolo D., Avolio R., Agliarulo I., Sarnataro D., Pacelli C., Scrima R., Colamatteo A., Matarese G., Capitanio N., Landriscina M, & Esposito F. (2022). Regulation of mitochondrial complex III activity and assembly by TRAP1 in cancer cells. Cancer Cell International, 22, 402.

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