Restriction enzymes, recombinant Taq DNA polymerase, and Pfu DNA polymerase were purchased from Fermentas (Milan, Italy). Ampicillin, chloramphenicol, isopropyl-β-d -thiogalactopyranoside (IPTG), 5-bromo-4-chloro-indolyl-β-d -galactopyranoside (X-Gal), and soluble starch were purchased from Sigma-Aldrich (USA). All chemicals were reagent grade. All oligonucleotide primers used in this study were synthesized by Sigma (Milan, Italy).
5 bromo 4 chloro indolyl β d galactopyranoside x gal
Manufactured by Merck Group
Sourced in United States
5-bromo-4-chloro-indolyl-β-d-galactopyranoside (X-Gal) is a synthetic organic compound used as a chromogenic substrate for the detection of β-galactosidase activity in biological systems. It is commonly employed in molecular biology and microbiology applications.
Automatically generated - may contain errors
Lab products found in correlation
Soluble starch, by Merck Group (1 mentions)
Restriction enzyme, by Thermo Fisher Scientific (1 mentions)
Ampicillin, by Merck Group (1 mentions)
Pfu dna polymerase, by Thermo Fisher Scientific (1 mentions)
Chloramphenicol, by Merck Group (1 mentions)
Isopropyl β d thiogalactopyranoside iptg, by Merck Group (1 mentions)
Recombinant taq dna polymerase, by Thermo Fisher Scientific (1 mentions)
Paraformaldehyde (pfa), by Merck Group (1 mentions)
Glutaraldehyde, by Merck Group (1 mentions)
Ethylene glycol tetraacetic acid (egta), by Merck Group (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Merck Group (1 mentions)
Np 40, by Merck Group (1 mentions)
Potassium ferricyanide, by Merck Group (1 mentions)
Mgcl2, by Merck Group (1 mentions)
Potassium ferrocyanide, by Merck Group (1 mentions)
Axio observer z1 microscope, by Zeiss (1 mentions)
Acetonitrile, by Merck Group (1 mentions)
Yeast extract, by Thermo Fisher Scientific (1 mentions)
Tryptone, by Thermo Fisher Scientific (1 mentions)
Dithiothreitol (dtt), by Sangon (1 mentions)
3 protocols using 5 bromo 4 chloro indolyl β d galactopyranoside x gal
1
Characterization of Recombinant Enzymes
2
Inner Ear Tissue Staining and Imaging
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Mice were euthanized and inner ears removed and fixed for 2 h at 4°C in 0.1 M phosphate buffer containing 1% paraformaldehyde (Sigma-Aldrich), 2 mM MgCl2 (Sigma-Aldrich), 0.25% glutaraldehyde (Sigma-Aldrich) and 5 mM EGTA (Merck Millipore). Ears were then washed in 0.1 M phosphate buffer containing 2 mM MgCl2 (Sigma-Aldrich) and 0.02% NP-40 (Fluka). Staining was performed overnight at room temperature (RT) in a solution of 0.1 M phosphate buffer containing 2 mM MgCl2 (Sigma-Aldrich), 5 mM potassium ferrocyanide (Sigma-Aldrich), 5 mM potassium ferricyanide (Sigma-Aldrich), 0.02% NP-40 and 1 mg/ml 5-bromo-4-chloro-indolyl-β-D-galactopyranoside (X-Gal) (Sigma-Aldrich). Post-staining, ears were decalcified in 4.3% EDTA in 0.1 M phosphate buffer (Sigma-Aldrich) for 48 h at 4°C, before paraffin embedding and sectioning at 10 µm. Sections and whole-mount dissected cochlea were imaged on a Zeiss Axio Observer Z-1 microscope using extended focus image capture.
3
Microbial β-Galactosidase Production and Characterization
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Medium components, such as tryptone and yeast extract, were purchased from Oxoid (Thermo Fisher Scientific, Waltahm, MA). Ortho-nitrophenyl-β-dgalactopyranoside (ONPG) and dithiothreitol were purchased from Sangon Biotech (Shanghai, China). Isopropyl-β-d-thiogalactopyranoside and 5-bromo-4chloro-indolyl-β-d-galactopyranoside (X-gal) were purchased from Sigma-Aldrich (Shanghai, China). Acetonitrile was purchased from Sigma-Aldrich. Molecular weight protein marker was purchased from Thermo Fisher Scientific. Protein determination kit was purchased from Zoman Biotechnology (Beijing, China). Glucose determination kit was purchased from Rongsheng Biotech (Shanghai, China). The restriction enzymes and polymerase were purchased from New England Biolabs (Beijing, China). Other reagents were of analytical reagent grade. The Escherichia coli hosts DH5α and BL21(DE3) were stocks in our laboratory. The commercial β-galactosidase derived from Aspergillus oryzae was purchased from Jiangsu Ruiyang Biotech Co. Ltd. (Wuxi, China). The milk used for the lactose hydrolysis experiment was fresh milk taken from the local dairy farm (Wuxi, China).
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