Restriction endonucleases, T4 DNA ligase, Phusion DNA polymerase, molecular mass markers and kits for DNA isolation were from Thermo Fisher Scientific (Waltham, MA, USA). LB (Luria-Bertani) medium was from Amresco (Solon, OH, USA), Tryptic Soy Broth, Brain Heart Infusion and Yeast Extract were from Difco-Becton Dickinson and Co. (Franklin Lakes, NJ, USA), general laboratory reagents were from Sigma-Aldrich (Moscow, Russia), liquid chromatography media were from GE Healthcare (Moscow, Russia), reagents for Western blot were from Bio-Rad (Moscow, Russia). The commercially available reagent “Absorbed staphylococcal anatoxin” (Medgamal, Moscow, Russia) was used as a crude immunogen. This reagent represents cell-free broth culture of S. aureus O15 inactivated by 0.4% formalin, concentrated by sequential treatment with trichloro acetic acid at pH=3.5 and 70% ethanol and absorbed on aluminium hydroxide. The same preparation but without formalin treatment and aluminium hydroxide absorption was used in Western blot and protein electrophoresis as “staphylococcal anatoxin” (i.e. non-absorbed).
Molecular mass marker
Manufactured by Thermo Fisher Scientific
Sourced in United States
Molecular mass markers are laboratory tools used to determine the molecular weight or mass of proteins or other macromolecules. They provide a reference point for comparison during electrophoresis experiments.
Automatically generated - may contain errors
Lab products found in correlation
T4 dna ligase, by Thermo Fisher Scientific (4 mentions)
Liquid chromatography media, by GE Healthcare (2 mentions)
Restriction endonuclease, by Thermo Fisher Scientific (2 mentions)
Kits for dna isolation, by Thermo Fisher Scientific (2 mentions)
Phusion dna polymerase, by Thermo Fisher Scientific (2 mentions)
General laboratory reagents, by Merck Group (1 mentions)
Reagents for western blot, by Bio-Rad (1 mentions)
Coomassie blue r 250, by HiMedia (1 mentions)
Pegfp c, by Takara Bio (1 mentions)
Pgex 4t 1, by GE Healthcare (1 mentions)
Pmal c5x, by New England Biolabs (1 mentions)
Anti flag antibody, by Cell Signaling Technology (1 mentions)
Coomassie brilliant blue r 250 stain, by Bio-Rad (1 mentions)
Mini protean, by Bio-Rad (1 mentions)
Amicon ultra, by Merck Group (1 mentions)
6 protocols using molecular mass marker
1
Staphylococcal anatoxin production
2
Evaluating Mucin Type-1S Purity
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Mucin type-1S purity was evaluated with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) using 8% polyacrylamide gel taking mucin in 1× PBS at the concentration of 50 mg mL−1. Then we diluted the mucin protein for SDS-PAGE gel run at the concentration of 30 µg and loaded 30 µL of mucin along with another protein, as experimental control, collagen, 30 µg of concentration with high molecular weight protein ladder. Molecular mass markers were purchased from (Thermo fisher scientific, # cat no. LC 5688) and the gel was stained with Coomassie blue R-250 (Himedia, cat no. MB153) using the manufacture's protocol.
3
Bacterial Expression of Recombinant Proteins
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Restriction endonucleases, T4 DNA ligase, Phusion DNA polymerase, molecular mass markers, and kits for DNA isolation were obtained from Thermo Fisher Scientific (Moscow, Russia). The lysogeny broth Miller recipe (LB) medium was obtained from Amresco (Solon, OH, USA); liquid chromatography media from GE Healthcare (Moscow, Russia); reagents for agarose and polyacrylamide gel electrophoresis from Bio-Rad (Moscow, Russia); and general laboratory reagents from Merck (Moscow, Russia).
Gene cloning and recombinant protein production were carried out in Escherichia coli DH10B and Rosetta (DE3), respectively (Merck). Vectors for cloning and recombinant protein expression in E. coli included pMal-c5x (cloning with the NH2-terminal maltose-binding protein (MBP), New England Biolabs, Ipswich, MA, USA) and pGEX4T-1 (cloning with the NH2-terminal glutathione-S-transferase (GST), GE Healthcare). Anti-MBP antibody (#E8032S) was purchased from New England Biolabs, and anti-FLAG antibody (#14793) from Cell Signaling (Leiden, The Netherlands). The source of the green fluorescent protein coding sequence (eGFP) was the plasmid pEGFP-C, obtained from Clontech (Takara Bio Europe SAS, Saint-Germain-en-Laye, France).
Gene cloning and recombinant protein production were carried out in Escherichia coli DH10B and Rosetta (DE3), respectively (Merck). Vectors for cloning and recombinant protein expression in E. coli included pMal-c5x (cloning with the NH2-terminal maltose-binding protein (MBP), New England Biolabs, Ipswich, MA, USA) and pGEX4T-1 (cloning with the NH2-terminal glutathione-S-transferase (GST), GE Healthcare). Anti-MBP antibody (#E8032S) was purchased from New England Biolabs, and anti-FLAG antibody (#14793) from Cell Signaling (Leiden, The Netherlands). The source of the green fluorescent protein coding sequence (eGFP) was the plasmid pEGFP-C, obtained from Clontech (Takara Bio Europe SAS, Saint-Germain-en-Laye, France).
4
Mealworm Protein SDS-PAGE Analysis
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For SDS-PAGE analyses, the alkaline and acidic mealworm protein extracts and digests were diluted 1:1 with Laemmli buffer (90 mM Tris-HCl pH 6.8, 4% (w/v) SDS, 20% (v/v) glycerol, 0.04% (w/v) bromphenol blue) with or without 10% β-mercaptoethanol, boiled for 5 min at 95 °C and separated on 12% acrylamide/Tris-HCl gels (MiniProtean, Bio-Rad, Hercules, CA, USA). Molecular mass markers (10–170 kDa; Thermo Fisher Scientific, Vilnius, Lithuania) were run in parallel to the samples. Following electrophoresis, the gels were either stained using Coomassie Brilliant Blue R-250 stain (Bio-Rad, München, Germany) to visualize protein components, or used for Western blotting.
5
Isoenzyme Patterns of Peroxidase and Superoxide Dismutase
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The detection of the isoenzyme patterns for two different groups of enzymes was performed on a native poly-acrylamide gel electrophoresis in the presence of low amounts of SDS (0.1%) in the running buffer. No reducing agents were present [54 (link)]. For the detection of the enzyme activities, the electrophoresis was carried out at 4 °C. Total proteins were separated and the resulting patterns were stained for peroxidase (POX) and superoxide dismutase (SOD) activities and total protein (Coomassie). Equal amounts of protein (50 μg) were loaded in each lane of the gels. The resolving gel and stacking gel had acrylamide concentrations of 10% and 5%, respectively. A molecular mass marker (Thermo Fisher Scientific, Dreieich, Germany) was run on all gels along with the tested samples.
6
SDS-PAGE Analysis of Soy and Lupine Proteins
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The fraction separated by FPLC method was subjected to electrophoresis in 14% polyacrylamide gels under denaturing conditions [18 (link)]. Samples obtained from soy chromatography were directly denatured, while lupine samples were first concentrated 10 times through filters with point cut-off 3 kDa (Amicon Ultra, Millipore Ltd.). Gels were dyed with Coomassie Brilliant Blue and documented using CLIQS (TotalLabQuant, GB). Molecular mass of the detected protein was determined by reference to molecular mass marker in range 20–120 kDa (Thermo Fisher Scientific, Waltham, USA).
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