For β-galactosidase assays, cells carrying pCSR2-lacZ plasmid (pSL168) were grown overnight in SC-based glucose medium to exponential phase. The A600 of each culture was measured and, for each assay, 1 ml of cells were harvested by centrifugation. The remaining cells were resuspended in SC-based lactate medium and grown for 2 h before the next assay. For each assay, cell pellets were frozen in liquid nitrogen and resuspended in 800 µl buffer Z (50 mM NaH2PO4, 45 mM Na2HPO4, 10 mM KCl, and 10 mM MgSO4, pH 7) to which β-mercaptoethanol was added (27 µl/10 ml; Sigma-Aldrich). Then, 160 µl ortho-nitrophenyl-β-galactoside at 4 mg/ml (Sigma-Aldrich) were added, and samples were incubated at 37°C for 30 min, which was still in the linear phase of the reaction. The reactions were stopped by adding 400 µl 1 M Na2CO3. The samples were then centrifuged for 2 min at 14,000 rpm to remove cell debris, and absorbance at 420 nm (A420) was measured. To remove background in each experiment, assays were performed similarly using cells that did not express the lacZ reporter gene. β-Galactosidase activities were calculated as 1,000 × [A420/(A600 × time)], where A420 is the mean of three technical replicates. For each experiment, histograms show the result and standard error of at least three independent biological replicates.
Ortho nitrophenyl β galactoside
Manufactured by Merck Group
Sourced in United States
Ortho-nitrophenyl-β-galactoside is a synthetic substrate used in biochemical and molecular biology applications. It is a colorless compound that undergoes a color change when cleaved by the enzyme β-galactosidase, making it useful as a reporter molecule.
Automatically generated - may contain errors
Lab products found in correlation
Dna purification kit, by Qiagen (2 mentions)
Ni nta resin, by Thermo Fisher Scientific (2 mentions)
Restriction endonuclease, by New England Biolabs (2 mentions)
Dna polymerase, by New England Biolabs (2 mentions)
T4 ligase, by New England Biolabs (2 mentions)
1 14c oleic acid, by American Radiolabeled Chemicals (2 mentions)
Oligonucleotide primers, by Integrated DNA Technologies (2 mentions)
β mercaptoethanol, by Merck Group (1 mentions)
Sodium acetate, by Merck Group (1 mentions)
Butylated hydroxytoluene bht, by Merck Group (1 mentions)
Ferrous sulfate heptahydrate, by Merck Group (1 mentions)
Trichloroacetic acid, by Merck Group (1 mentions)
2 2 diphenyl 1 picrylhydrazyl (dpph), by Merck Group (1 mentions)
Ethyl alcohol, by Merck Group (1 mentions)
Quercetin, by Merck Group (1 mentions)
Aluminum chloride, by Merck Group (1 mentions)
2 2 azino bis 3 ethylbenzothiazoline 6 sulfonic acid diammonium salt, by Merck Group (1 mentions)
Phenazine methosulfate, by Merck Group (1 mentions)
Ferrous chloride, by Merck Group (1 mentions)
Phenol reagent, by Merck Group (1 mentions)
7 protocols using ortho nitrophenyl β galactoside
1
β-Galactosidase Assay Protocol
2
Measuring SOS Response in S. aureus
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The induction of SOS response was measured using a derivative S. aureus 8325-4 strain carrying a recA::lacZ fusion. Bacteria were grown exponentially in LB medium until an OD600 between 0.1 and 0.2. Cells were treated with SteLL or ciprofloxacin (both at 0.5 × MIC) for 3 h70 (link). Cells were permeabilizated by toluene and β-galactosidase activity was measured using ONPG (ortho-Nitrophenyl-β-galactoside; Sigma-Aldrich).
3
Antioxidant Capacity Assay Collection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Ethyl alcohol (95%), ortho-nitrophenyl-β-galactoside, Folin–Ciocalteu's phenol reagent, Griess' reagent for nitrite, aluminum chloride, β-nicotinamide adenine dinucleotide hydrate (NADH), phenazine methosulfate (PMS), 2,4,6-tris (2-pyridyl)-s-triazine (TPTZ), sodium acetate, L-ascorbic acid, quercetin, gallic acid, ferrous chloride, ferrous sulfate heptahydrate, trichloroacetic acid (TCA), 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), (±)-6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid (Trolox), potassium persulfate, sodium nitroprusside, butylated hydroxytoluene (BHT) were purchased from Sigma Aldrich (St. Louis, MO, USA).
4
Yeast FLO11 Gene Expression Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Plasmids pLG669-Z FLO11-6/7 and -9/10 were transformed into designated yeast strains by standard protocols [64] . Cells were grown in SC-Ura media overnight, and then inoculated into fresh media to an OD600 of approximately 0.2. Cultures were subsequently grown for 3–4 hours to an OD600 of approximately 0.8–1.0. β-galactosidase activity was determined with the use of ortho-nitrophenyl-β-galactoside as a substrate (Sigma Aldrich) [70] (link), [71] (link).
5
Molecular Cloning Reagents and Protocols
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Kanamycin, lysozyme, and ortho-Nitrophenyl-β-galactoside (ONPG) were purchased from Sigma-Aldrich Corporation (St. Louis, MO). Ampicillin and bovine serum albumin (BSA) were bought from Fisher Scientific (Fairlawn, NJ). Isopropyl-β-D-thiogalactopyranoside (IPTG) was obtained from Anatrace, Inc (Maumee, Ohio). All restriction enzymes, T4 DNA ligase, and T4 polynucleotide kinase were purchased from New England Biolabs (Beverly, MA). Pfu DNA polymerase was obtained from Stratagene, Inc. (La Jolla, CA). All synthetic oligonucleotides were purchased from Eurofins Genomics (Huntsville, AL). Plasmid and DNA fragment cleaning kits including QIAprep Spin Miniprep Kit, QIAquick Gel Extraction Kit, and QIAquick Nucleotide Removal Kit were obtained from QIAGEN, Inc. (Valencia, CA). Luciferase Assay System was bought from Promega Corporation (Madison, WI).
6
Synthesis and Analysis of Radiolabeled Fatty Acids
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The cis-9, 10-ethyleneoctadecanoic acid was purchased from Cayman Chemicals and all of the other fatty acids, antibiotics, and ortho-nitrophenyl-β-galactoside (ONPG) were purchased from Sigma-Aldrich. DNA polymerases, restriction endonucleases, and T4 ligase were from NEB and the TA cloning kit was from Thermo-Fisher Scientific. Sodium [1-14C]acetate (specific activity, 58.6 mCi/mmol) and [1-14C]stearic acid (specific activity, 53.0 mCi/mmol) were provided by Moravek, Inc while the [1-14C]oleic acid (specific activity, 55 mCi/mmol) was purchased from American Radiolabeled Chemicals. Ni-NTA resin, 6% native PAGE gel and SYBR Green I nucleic acid stain were from Invitrogen and the DEAE anion-exchange column was from Sartorius. The DNA purification kits were from Qiagen, and the silver nitrate silica gel thin layer plates were from Analtech. All the other reagents were of the highest available quality. Oligonucleotide primers were synthesized by Integrated DNA Technologies and DNA sequencing was provided by ACGT Inc.
7
Enzymatic Reactions with Radiolabeled Fatty Acids
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The 15-methyl palmitic acid was purchased from Cayman Chemicals and all the other fatty acids, antibiotics, phospholipase A2, ortho-nitrophenyl-β-galactoside, and agmatine sulfate were purchased from Sigma-Aldrich. The media were purchased from Fisher Scientific. The DNA polymerase, restriction endonuclease, and T4 ligase were purchased from New England Biolabs. Sodium[1-14C]acetate (specific activity, 57.0 mCi/mmol) and [1-14C]stearic acid (specific activity, 53.0 mCi/mmol) were provided by Moravek, Inc, and the [1-14C]oleic acid (specific activity, 55 mCi/mmol) was purchased from American Radiolabeled Chemicals. Ni-NTA resin was from Invitrogen, and the DNA purification kits were from Qiagen. Silver nitrate silica gel thin layer plates were purchased from Analtech and M17 Broth was from Becton Dickenson. All the other reagents were of the highest available quality. Oligonucleotide primers were synthesized by Integrated DNA Technologies, and DNA sequencing was performed by ACGT, Inc.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!