Rnase a

Manufactured by Promega

Sourced in United States, Germany, France, Australia

RNase A is a purified enzyme that catalyzes the hydrolytic cleavage of single-stranded RNA. It is commonly used in molecular biology workflows to degrade RNA.

Automatically generated - may contain errors

Lab products found in correlation

Propidium iodide, by Merck Group (11 mentions) Proteinase k, by Merck Group (4 mentions) Dnase 1, by Thermo Fisher Scientific (4 mentions) Propidium iodide, by Thermo Fisher Scientific (4 mentions) Facscalibur, by BD (3 mentions) Dnase, by Promega (3 mentions) T7 rna polymerase, by Takara Bio (3 mentions) Proteinase k, by New England Biolabs (3 mentions) Ethylene diamine tetraacetic acid (edta), by Merck Group (3 mentions) Propidium iodide solution, by Merck Group (2 mentions) Proteinase k, by Promega (2 mentions) Penicillin, by Thermo Fisher Scientific (2 mentions) Streptomycin, by Thermo Fisher Scientific (2 mentions) T7 rna polymerase ver 2, by Takara Bio (2 mentions) Rabbit reticulocyte lysate translation system, by Promega (2 mentions) Rnaclean xp, by Beckman Coulter (2 mentions) Ddctp, by Thermo Fisher Scientific (2 mentions) Easy a cloning kit, by Agilent Technologies (2 mentions) Tdt buffer, by Promega (2 mentions) Nextseq platform, by Illumina (2 mentions)

Spelling variants of this product

RNase A solution (13 citations) DNase-free RNase A Solution (8 citations)

82 protocols using rnase a

Proteomics Protocol for Enriching RNA-Peptides

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Starting from the methanol-precipitated OOPS interface, proteins were digested using 1 μg Lys-C (Promega, Madison, WI, USA) in 100 μL of 100 mM TEAB (Sigma-Aldrich) with 1 μL of RNaseOUT (Thermo Fisher Scientific) overnight at 37 ºC. Two different approaches were used to enriched RNA-peptides:
RNA-peptides were re-suspended in 100 μL of 100 mM Tris-HCl (pH 8.0)/ 2 mM MgCl₂, sonicated for 15 min and incubated at 95 ºC for 20 min. 2 μg RNase A/T1 mix (2 mg/mL of RNase A and 5000 U/mL of RNase T1) was added to cooled samples, and incubated for 4 h at 37 ºC followed by a second protease digestion using 1 μg trypsin (Promega) overnight at 37 ºC. Digested samples were desalted with Oligo R3 as described in the “proteomics sample preparation” section and dried on speedvac (Labconco).
Digests were re-suspended in 30-40 μL of 80% acetonitrile (ACN)/2% TFA containing 1 μg of TiO₂ beads (GL Sciences). The slurry was transferred into a p200 tip containing a C8 “plug” (3M Empore, Sigma-Aldrich) to retain the loaded TiO₂ beads and the flow-through collected. The packed TiO₂ was washed with 20 μL 80% ACN/2% TFA, then 20 μL 10% ACN/0.1% TFA and the flow-through from both retained. The TiO₂-enriched fraction was eluted from the beads with two rounds of 20 μL of ammonia solution (1.5-1.8%), pH>10.5, and 20 μL of 50% ACN.

Queiroz R.M., Smith T., Villanueva E., Marti-Solano M., Monti M., Pizzinga M., Mirea D.M., Ramakrishna M., Harvey R.F., Dezi V., Thomas G.H., Willis A.E, & Lilley K.S. (2019). Comprehensive quantitation of RNA-protein interaction dynamics by orthogonal organic phase separation (OOPS). Nature biotechnology, 37(2), 169-178.

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DATS Inhibits HNSCC Cell Growth

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Cell culture essentials, including DMEM (Dulbecco’s Modified Eagle Medium), sodium pyruvate, non-essential amino acids, fetal bovine serum (FBS), and penicillin/streptomycin antibiotic mixture, were obtained from GIBCO (Grand Island, NY, USA). Diallyl trisulfide (DATS, purity > 98%) was procured from LKT Laboratories (St. Paul, MN, USA), and RNase A was sourced from Promega (Madison, WI, USA). Cyclin B1, Cdk1, pERK1/2, ERK1 antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA); the anti-beta actin antibody was purchased from Sigma (St. Louis, MO, USA); and the antibody against cleaved poly-(ADP-ribose)-polymerase (c-PARP), cleaved caspase-3, pAkt (Ser 473), Akt, γH2AX (Ser 139), were purchased from Cell Signaling Technology (Danvers, MA, USA). UMSCC-22A, UMSCC-22B, and Cal33 cells were a generous gift from Prof. Daniel E. Johnson (University of California, San Francisco, CA, USA). HNSCC cells were grown in DMEM and supplemented with 10% FBS, 100 µg/mL streptomycin, and 100 units/mL penicillin under standard culture conditions. Stock solution of DATS was prepared in dimethyl sulfoxide (DMSO).

Mathan S.V., Singh R., Kim S.H., Singh S.V, & Singh R.P. (2024). Diallyl Trisulfide Induces ROS-Mediated Mitotic Arrest and Apoptosis and Inhibits HNSCC Tumor Growth and Cancer Stemness. Cancers, 16(2), 378.

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DNA Fragmentation Assay in 6-OHDA-Treated MN9D Cells

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To assess DNA fragmentation, MN9D cells were treated with 100 μM 6-OHDA for 18 h. Soluble DNA was harvested by lysing cells for 30 min at 50 °C in buffer consisting of 0.5% Triton X-100, 5 mM Tris-HCl (pH 7.4), 20 mM EDTA, and 20 μg/mL proteinase K. Following microcentrifugation at 13,000 g at 4 °C for 15 min, DNA in the supernatant was extracted with phenol/chloroform and ethanol precipitated. DNA precipitates were dissolved in an appropriate amount of double-distilled H₂O and incubated with 4 mg/mL RNase A (Promega) for 1 h at 37 °C. Purified DNA (30 μg) was electrophoresed on 1.5% agarose gels and stained with ethidium bromide. Intercalated DNA bands were viewed on a UV transilluminator using a Molecular Imager Gel Dox XR system (Bio-Rad).

Oh C.K., Choi Y.K., Hwang I.Y., Ko Y.U., Chung I.K., Yun N, & Oh Y.J. (2020). RING-finger protein 166 plays a novel pro-apoptotic role in neurotoxin-induced neurodegeneration via ubiquitination of XIAP. Cell Death & Disease, 11(10), 939.

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Measuring Cell Proliferation and Cytotoxicity

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For measuring cellular proliferation, cells were seeded at 100000 cells per well in 6-well plates and counted using a hemocytometer 4 days later. The values of doubling times were obtained applying the exponential growth formula and further used for calculation of doublings per day as previously reported.^{21 (link)} To measure cytotoxicity, trypan blue (Sigma)-positive cells were counted. For cell cycle analysis, cells were harvested and fixed with 70% ethanol overnight at 4°C. The following day the cells were washed twice with PBS and incubated with 40 µg/mL propidium iodide solution (Sigma) containing 100 µg/mL RNase A (Promega) for 30 minutes at 37°C. The cell cycle phase was determined by flow cytometry (LSR-Fortessa, BD Biosystems) and analyzed using FlowJo software.

Park J.W., Wollmann G., Urbiola C., Fogli B., Florio T., Geley S, & Klimaschewski L. (2018). Sprouty2 enhances the tumorigenic potential of glioblastoma cells. Neuro-Oncology, 20(8), 1044-1054.

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Cell Cycle Analysis After RF Exposure

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The cells after 169MHz RF exposure for different times (0, 30, 90, 180 and 1440 mins) and relative positive control exposure were collected and fixed in 70% ethanol at −20°C, overnight (O.N.), three washed with 1X PBS, and finally dissolved in a hypotonic buffer containing propidium iodide (1XPBS), 100μg/mL of RNAse A (Promega, Madison, WI, USA) and 40μg/mL of Propidium iodine (Sigma-Aldrich) and incubated for 30mins at RT in dark. Samples were acquired on a Guava^® easyCyte™ flow cytometer (Millipore, Sigma) and analyzed using the standard procedure recommended by the easyCyte™ software.

Alessio N., Santoro E., Squillaro T., Aprile D., Briccola M., Giubbini P., Marchesani R., Muoio M.R, & Lamberti M. (2019). Low-Level Radiofrequency Exposure Does Not Induce Changes in MSC Biology: An in vitro Study for the Prevention of NIR-Related Damage. Stem Cells and Cloning : Advances and Applications, 12, 49-59.

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Apoptosis and Cell Cycle Analysis in Bladder Cancer

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The bladder cancer cells transfected with pcDNA-CASC2 or empty vector were harvested for 48 h for cell apoptosis and cell cycle analysis. The cell apoptosis was determined by the Annexin V-FITC apoptosis kit (Sigma-Aldrich Chemical Company, St Louis MO, USA) according to the manufacturer's instructions. The Annexin V-FITC and PI fluorescence levels were measured by flow cytometry (BD Biosciences, FACS Calibur). The Annexin V-positive cells (both PI-negative and -positive) were defined as apoptotic cells. For the cell cycle analysis, cells were collected and fixed with 70 % pre-cooling anhydrous ethanol at −20 °C overnight and incubated with propidium iodide (PI) dye liquor (Sigma) in the presence of Rnase A (Promega) for 30 min at room temperature. The cell cycle distribution was analyzed by flow cytometry (FACSCalibur, BD Biosciences).

Pei Z., Du X., Song Y., Fan L., Li F., Gao Y., Wu R., Chen Y., Li W., Zhou H., Yang Y, & Zeng J. (2017). Down-regulation of lncRNA CASC2 promotes cell proliferation and metastasis of bladder cancer by activation of the Wnt/β-catenin signaling pathway. Oncotarget, 8(11), 18145-18153.

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Extracellular Vesicle Isolation Protocol

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First, 40 mL of BCBL-1 cell culture supernatant was spun at 1000 g at 4°C for 10 min to pellet cells and large debris. Second, samples were transferred to clean, 50 mL conical tubes and spun at 16,000 g at 4°C for 30 min to remove larger vesicles such as apoptotic bodies. Third, clarified cell supernatant was passed through a 0.22 µm Nalgene Rapid Flow Filter as above. The clarified solution was placed into disposable plastic tubes and into the Beckman Optima LE-80K Ultracentrifuge using the SW32-TI rotor at 100,000 g for 60 min at 4°C at < 20μ. Excess fluid was aspirated, the pellet was washed with 5 mL of 1X PBS and spun again at 100,000 g for 60 min at 4°C at < 20μ. The first UC pellet was resuspended in 250 μL of PBS, and the solution was then incubated with RNaseA (Promega, P: A7973) at a final concentration of 50 µg/mL at 37°C for 30 min. The solution was then diluted to 5 mL and spun as before two more times (three total UC steps). The final pellet was resuspended in 50 µL of 1X PBS, yielding a concentration factor of ~ 800-fold (0.05 µL/40 ml).

McNamara R.P., Caro-Vegas C.P., Costantini L.M., Landis J.T., Griffith J.D., Damania B.A, & Dittmer D.P. (2018). Large-scale, cross-flow based isolation of highly pure and endocytosis-competent extracellular vesicles. Journal of Extracellular Vesicles, 7(1), 1541396.

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Extracellular Vesicle Fractionation and Effects

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ELV were purified from CCCM and ICCM, and then divided into four fractions. The first fraction was non-treated. The second, third, and fourth fractions were treated with RNase A (0.4 μg/μl), DNase (Promega), protease K (100 μg/ml), respectively, for 30 min in each case, and then each of the fractions was used to treat HDFn cells, as reported previously^{42 (link)–44}.

Ariyoshi K., Miura T., Kasai K., Fujishima Y., Nakata A, & Yoshida M. (2019). Radiation-Induced Bystander Effect is Mediated by Mitochondrial DNA in Exosome-Like Vesicles. Scientific Reports, 9, 9103.

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Stability of M and HA Genes under RNase

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To assess the stability of M and HA genes under direct action of RNase, the RNA of the inactivated viral strain A/chicken/Egypt/1709-6/2008 was incubated with RNase A dilution (Promega, Madison, WI, USA). Thus, 150 µL of viral RNA were mixed with 4.5 µL of RNase A dilution (4 mg/mL diluted 1:1000) and incubated at 37 °C for different time periods (0, 30 s, 1 min, 2 min, 3 min, 4 min, 5 min, 6 min and 7 min). The integrity of the diagnostic target regions was assessed by qRT-PCR.

Relova D., Rios L., Acevedo A.M., Coronado L., Perera C.L, & Pérez L.J. (2018). Impact of RNA Degradation on Viral Diagnosis: An Understated but Essential Step for the Successful Establishment of a Diagnosis Network. Veterinary Sciences, 5(1), 19.

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Apoptosis and Cell Cycle Analysis

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The cell apoptosis was determined by the Annexin V-FITC apoptosis kit (BD Biosciences, USA) according to the manufacturer’s guide. Briefly, cells were harvested and washed with PBS, stained in 500μl binding buffer with 5μl of Annexin V-FITC (BD Biosciences) for 15 min and 5μl PI for another 5 min. Apoptosis were measured by flow cytometry (BD Biosciences, FACS Calibur). For the cell cycle analysis, cells were harvested and washed with cold PBS, fixed with 75 % ice-cold ethanol at −20°C overnight, stained with propidium iodide (PI) dye solution (Sigma) supplemented 0.1 mg/ml Rnase A (Promega) for 30 min at room temperature. The cell cycle distribution was analyzed by flow cytometry (FACSCalibur, BD Biosciences).

Zhu Y., Li B., Liu Z., Jiang L., Wang G., Lv M, & Li D. (2017). Up-regulation of lncRNA SNHG1 indicates poor prognosis and promotes cell proliferation and metastasis of colorectal cancer by activation of the Wnt/β-catenin signaling pathway. Oncotarget, 8(67), 111715-111727.

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