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RNase is an enzyme that degrades ribonucleic acid (RNA). It is commonly used in molecular biology and biochemistry laboratories to remove unwanted RNA from samples.

Automatically generated - may contain errors

Lab products found in correlation

Genejet plant rna purification kit, by Thermo Fisher Scientific (2 mentions) Primescript rt pcr kit, by Takara Bio (2 mentions) Tb green premix ex taq, by Takara Bio (2 mentions) Pe conjugated mouse igg2b, by R&D Systems (1 mentions) Mouse igg2b, by Merck Group (1 mentions) Viia 7 real time pcr system, by Thermo Fisher Scientific (1 mentions) Mirvana paris total rna isolation kit, by Thermo Fisher Scientific (1 mentions) Interleukin 6 (il 6), by Thermo Fisher Scientific (1 mentions) Arginase 1, by Thermo Fisher Scientific (1 mentions) Superscript vilo cdna synthesis kit, by Thermo Fisher Scientific (1 mentions) Gapdh, by Thermo Fisher Scientific (1 mentions)

4 protocols using rdnase

1

Quantifying DFR and PAL2 Gene Expression

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Total RNA was isolated from 100 mg of fresh leaf tissue harvested at 5 dpi by using the GeneJET Plant RNA Purification Kit (Thermo Fisher Scientific). The RNA was treated with rDNase (Invitrogen) before cDNA synthesis. The cDNA was synthesized from 1 µg of total RNA using the PrimeScript™ RT-PCR Kit (Takara). Three technical replicates were measured for gene expression levels in each cDNA sample. The SYBR® fluorescent dye (TB Green® Premix Ex Taq™, Takara) was employed for reporting the amplification of cDNA in the Applied biosystem 7500 Fast Real-Time PCR system with specific primer pairs for either DFR or PAL2 genes (Additional file 1: Table S2). F-BOX gene was amplified as an internal reference (Liu et al., 2012) for determining DFR/PAL2 gene expression in each sample (ΔCt). The differential DFR/PAL2 gene expression levels were calculated according to the comparative ΔΔCt method as the ratio between the samples agroinfiltrated with the dCasEV2.1 and a gRNA targeting the DFR/PAL2 promoter (gNbDFR/gNbPAL2) and those including the same construct but an unrelated gRNA (gMTB), both for copper and water treatments [19 ].
Garcia-Perez E., Diego-Martin B., Quijano-Rubio A., Moreno-Giménez E., Selma S., Orzaez D, & Vazquez-Vilar M. (2022). A copper switch for inducing CRISPR/Cas9-based transcriptional activation tightly regulates gene expression in Nicotiana benthamiana. BMC Biotechnology, 22, 12.
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2

Quantifying DFR Gene Expression

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Total RNA was isolated from 100 mg of fresh leaf tissue harvested at 5 dpi by using the GeneJET Plant RNA Purification Kit (Thermo Fisher Scientific). The RNA was treated with rDNase (Invitrogen) before cDNA synthesis. The cDNA was synthesized from 1 µg of total RNA using the PrimeScript™ RT-PCR Kit (Takara). Three technical replicates were measured for gene expression levels in each cDNA sample. The SYBR ® fluorescent dye (TB Green ® Premix Ex Taq™, Takara) was employed for reporting the amplification of cDNA in the Applied biosystem 7500 Fast Real-Time PCR system with specific primer pairs for DFR gene (Supplementary Table 2). F-BOX gene was amplified as an internal reference (Liu et al., 2012) for determining DFR gene expression in each sample (ΔCt). The differential DFR gene expression levels were calculated according to the comparative ΔΔCt method as the ratio between the samples agroinfiltrated with the dCasEV2.1 and a gRNA targeting it to the DFR promoter (gDFR) and those including the same construct but an unrelated gRNA (gMTB), both for copper and water treatments (Livak & Schmittgen, 2001) (link).
Garcia-Perez E., Diego‐Martin B., Quijano‐Rubio A., Giménez E.M., Orzáez D., & Vázquez‐Vilar M. (2021). A copper switch for inducing CRISPR/Cas9-based transcriptional activation tightly regulates gene expression in Nicotiana benthamiana.
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3

Quantification and Characterization of Extracellular Vesicles

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MPs were purified from treated cell supernatants as described above. After suspension of MPs in PBS, the MP numbers were quantitated by flow cytometry as described above. For experiments to digest DNA, MPs were treated with recombinant DNase I (rDNase; 10U rDNase / 4×106 MPs; Life Technologies) for 60 min at 37°C. After this treatment, the MPs were transferred to flow cytometry tubes at an MP density of 2×105/tube. To assess HMGB1 content, samples were stained with a Phycoerythrin (PE)-conjugated mouse anti-human HMGB1 antibody (2ul/tube; monoclonal antibody directed to residues 2–215, R&D systems, Minneapolis, MN) or a PE-conjugated mouse IgG2b (R&D Systems) for 60min at 20°C. To assess DNA, MPs were stained with a mouse monoclonal anti-dsDNA antibody (clone 163p.132; 4ug/tube; a kind gift from Dr. Tony Marion) or a mouse IgG2b (isotype control; 4ug/tube; Sigma-Aldrich). After this step, MPs treated with either anti-DNA or isotype control were incubated with a F(ab')2 sheep anti-mouse IgG PE (Sigma-Aldrich) for 30min at 20°C in the dark. The samples were then analyzed by flow cytometry.
Spencer D.M., Mobarrez F., Wallén H, & Pisetsky D.S. (2014). The Expression of HMGB1 on Microparticles from Jurkat and HL-60 Cells Undergoing Apoptosis in vitro. Scandinavian journal of immunology, 80(2), 101-110.
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4

Quantifying Immune Markers in Adipose Tissue

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Frozen abdominal or pericardial fat tissues (~50mg) were homogenized in 350ul of ice cold lysis buffer, supplied by mirVana PARIS total RNA isolation kit (Life Technologies, Cat# AM1556).
Total RNA was then isolated from homogenized samples, and its concentration measured by a Spectrophotometer (NanoDrop). 50μl of the RNA samples were then treated with 2 units for rDNase (Life Technologies, Cat# AM1906) to remove possible contaminating DNA. First-strand cDNA was produced from 300ng of total RNA using SuperScript VILO cDNA Synthesis kit (Life Technologies, Cat#11754-050). Relative quantitative PCR was performed using Taqman assays, containing 20ng of cDNA products. Taqman assays included the followings: iNOS (Ss03374608), CD86 (Ss03394398), mannose-receptor (Ss03373693), CD11c (AJ1RVEU), TNF-a (Ss03391318), IL-6 (Ss03384604), CD-68 (AJWR2PY), arginase-1 (Ss03391398) and GAPDH (Ss03374854, for internal control), all from Life Technologies. Negative controls with no cDNA were cycled in parallel with each run. PCR analysis was done on Applied Biosystems ViiA7 Real-Time PCR systems at the following conditions: 50°C for 2 minutes, 95°C for 10 minutes and 40 cycles of 95°C for 15 seconds and 60°C for 1 minute. Fold-changes of each target gene in the obese relative to lean group were calculated using the 2-ΔΔCT method. Fold changes greater than 2 are considered as significant.
Pawar A.S., Zhu X.Y., Eirin A., Tang H., Jordan K.L., Woollard J.R., Lerman A, & Lerman L.O. (2014). Adipose tissue remodeling in a novel domestic porcine model of diet-induced obesity. Obesity (Silver Spring, Md.), 23(2), 399-407.
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