Dnase 1

Manufactured by Corning

Sourced in United States

DNAse I is an enzyme that catalyzes the hydrolytic cleavage of DNA. It is commonly used in molecular biology and biochemistry applications to degrade DNA.

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Lab products found in correlation

Dnase 1, by Merck Group (2 mentions) Tissue culture flask, by Corning (1 mentions) Dmem f12 medium, by Merck Group (1 mentions) Trypsin, by Worthington (1 mentions) Elastase, by Worthington (1 mentions) Collagenase type 1, by Thermo Fisher Scientific (1 mentions) Antibiotic antimycotic solution, by Corning (1 mentions) Dihydrotestosterone dht, by Merck Group (1 mentions) Dnase 1, by Roche (1 mentions) Facscanto 2, by BD (1 mentions) Tryple express, by Thermo Fisher Scientific (1 mentions) Dnase 1, by Worthington (1 mentions) Liberase tl, by Roche (1 mentions)

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DNase (2 citations)

5 protocols using dnase 1

Isolation of Alveolar Type I Cells

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After the removal of visible bronchi, the lung tissue was minced into pieces of thickness <0.5 mm with scissors. Lung pieces were washed with Hanks’s balanced salt solution at pH7.4 for three times to remove macrophages and blood cells. A combination of 0.5% trypsin and 4 U/ml elastase (Worthington Biochemical Corporation, USA) was added to the lung pieces and incubated for 40 min in a 37 °C water bath with shaking for digestion and stopped with DMEM/F12 medium with 40%FBS and DNase I (350 U/ml) (Sigma, USA). Undigested lung fragments were separated with a 50 μm pore size disposable cell strainer and cell clumps were dispersed by repeated pipetting for 10 min. Single cell suspension in flow-through was centrifuged and resuspended in a 1:1 mixture of DMEM/F12 medium and small airway growth medium (SAGM) (Lonza, USA) with 5% FBS and 350 U/ml DNase I. Resuspended cells were plated on a tissue culture flask (Corning, USA) for 90 min adhesion at 37 °C. The non-adherent cells were centrifuged, pelleted and resuspended with SAGM with supplements and 1%PS and plated on a new culture flask. The growth medium was changed daily starting from 60 h after cell plating. At 75% confluence, the ATI cell layer was trypsinised for seeding.

Chan L.L., Bui C.T., Mok C.K., Ng M.M., Nicholls J.M., Peiris J.S., Chan M.C, & Chan R.W. (2016). Evaluation of the human adaptation of influenza A/H7N9 virus in PB2 protein using human and swine respiratory tract explant cultures. Scientific Reports, 6, 35401.

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Tissue Digestion and Cell Isolation Protocol

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Fresh tissue samples were transported in ice-cold saline and immediately dissected into portions for fixation in 10% formalin followed by paraffin embedding. For human specimens, a 4-hour enzymatic digestion into single cells was performed at 37°C in 35 mL Hanks’ balanced salt solution (HBSS) containing 5 mg/mL Collagenase Type I (Life Technologies), 10 μM ROCK Inhibitor Y-27632 (StemRD), 1 nM dihydrotestosterone (DHT) (Sigma-Aldrich), 1 mg DNAse I (Roche), and 1% antibiotic/antimycotic solution (100×; Corning).^{4 (link)} Mouse specimens were digested for 1 hour in HBSS containing either 10 mg/mL cold protease or 1.5 to 2 mg/mL Collagenase Type I, plus 10 μM ROCK Inhibitor Y-27632, 1 nM DHT, 1 mg DNAse I, and 1% antibiotic/antimycotic solution.

Joseph D.B., Henry G.H., Malewska A., Iqbal N.S., Ruetten H.M., Turco A.E., Abler L.L., Sandhu S.K., Cadena M.T., Malladi V.S., Reese J.C., Mauck R.J., Gahan J.C., Hutchinson R.C., Roehrborn C.G., Baker L.A., Vezina C.M, & Strand D.W. (2020). Urethral luminal epithelia are castration-insensitive cells of the proximal prostate. The Prostate, 80(11), 872-884.

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Profiling Tumor-Infiltrating Immune Cells

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Omentum tumors were harvested and placed in digestion buffer (DMEM 25 Mm HEPES, 3.5% fatty acid free BSA plus 0.5 mg/ml collagenase +70μg/ml DNAse I) (all from Corning) and fragmented with scissors. Tumor fragments were placed in an orbital shaker at 250–300 rpm for 30 min at 37 °C and subsequently filtered through 70 μm nylon mesh, washed with staining buffer (1× PBS, 2% BCS, and 2 mM EDTA) (all from Corning), and centrifuged at 800 xg for 10 min. The following antibodies were used for staining: CD3 (100214CD3), NK1.1 (108716) CD45.2 (109824), (all from Biolegend, San Diego, CA, USA), CD8a (551,162 A 1), CD5 (553023), CD19 (11–0191–85), zombie aqua (423101) (all from BD Biosciences, San Jose, CA, USA), CD4 (Q10092) (Invitrogen, Carlsbad, CA, USA) and live-dead discriminator 7-Amino-AMD (7AAD) (129935) (Calbiochem/Millipore Sigma, Burlington, MA, USA). Samples were run using a BD FACS Canto II (BD Biosciences), DIVA software 6.1.3. Data were analyzed with FlowJo version 10.9.6.

Wall J.A., Meza-Perez S., Scalise C.B., Katre A., Londoño A.I., Turbitt W.J., Randall T., Norian L.A, & Arend R.C. (2020). Manipulating the Wnt/β-catenin signaling pathway to promote anti-tumor immune infiltration into the TME to sensitize ovarian cancer to ICB therapy. Gynecologic oncology, 160(1), 285-294.

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Isolation of Prostate Cells from Tissue

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Prostate tissues were minced with scissors and then incubated in papain (20 units/ml) with 0.1 mg/ml DNase I (Worthington LK003150) at 37°C with gentle agitation. After 45 min, samples were gently triturated, then incubated for another 20–45 min in papain as needed. Samples were gently triturated again, followed by quenching of the enzyme using 1 mg/ml ovomucoid/bovine serum albumin solution with 0.1 mg/ml DNase I (Worthington LK003150). Cells were passed through a 70 µm strainer (pluriSelect 43-10070-70) and washed with PBS-EDTA with 0.1 mg/ml DNase I (Corning MT-46034CI). If needed, the samples were additionally digested in TrypLE Express (Invitrogen 12605–036) for 3–5 min at 37°C with gentle agitation. The samples were gently triturated and the TrypLE was inactivated by addition of HBSS with 10% FBS and 0.1 mg/ml DNase I. Samples were passed through a 20 μm strainer (pluriSelect 43-10020-70), washed in 1x PBS, and resuspended in appropriate buffers for downstream analyses.

Crowley L., Cambuli F., Aparicio L., Shibata M., Robinson B.D., Xuan S., Li W., Hibshoosh H., Loda M., Rabadan R, & Shen M.M. (2020). A single-cell atlas of the mouse and human prostate reveals heterogeneity and conservation of epithelial progenitors. eLife, 9, e59465.

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Isolation of Murine Immune Cells

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Spleen and Peyer’s patch cells were isolated by mechanical disruption of the tissue between frosted glass slides and filtration through 66 μm nitex mesh. BM cells were isolated by flushing bones using a 23G needle and syringe with FACs buffer (PBS, 0.5% BSA, 1mM EDTA). siLP preparations were made using a modified protocol from Hall et al. (Hall et al., 2011 (link)). Small intestines were harvested and Peyer’s patches, fat, and intestinal contents were removed. The cleaned small intestine was cut into ~2cm pieces before incubating at 37° C on a shaker for 20 minutes in RPMI 1640 with 20mM HEPES, 5mM EDTA, 1mM DTT, Penicillin/Streptomycin, and 5% FBS. The small intestine pieces are then filtered over a large pore strainer and washed 2 times in RPMI with Pen/Strep, 2mM EDTA, and 20 mM HEPES. The pieces were minced and put into RPMI with Pen/Strep, 0.1 mg/mL Liberase TL (Roche), 0.05% DNase I (Sigma D5025), and 20 mM HEPES for 30 minutes at 37° C while shaking. The Liberase reaction is then quenched with cold RPMI with 10% FBS, 0.05% DNase I, Pen/Strep, 20 mM HEPES and the small intestine is filtered through 70 μm Corning cell strainers (Corning 352350). Before running siLP cells through the cell sorter or flow cytometer and before plating on ELISpot plates they are filtered again with 66 μm nitex.

Wilmore J.R., Gaudette B.T., Atria D.G., Hashemi T., Jones D.D., Gardner C.A., Cole S.D., Misic A.M., Beiting D.P, & Allman D. (2018). Commensal microbes induce serum IgA responses that protect against polymicrobial sepsis. Cell host & microbe, 23(3), 302-311.e3.

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