The ability of producing extracellular enzymes was evaluated on chromogenic reaction medium containing different substrates: 0.5% CM-cellulose (Sigma-Aldrich, St. Louis, MO, USA), D-cellobiose (Sigma-Aldrich), polygalacturonic acid (MP Biomedicals, Sanata Ana, CA, USA), starch (Sigma-Aldrich), xylan (Sigma-Aldrich), and avicel cellulose (Fluka, Cork, Ireland) as a carbon source, 0.1% yeast nitrogen base without amino acid (BD Biosciences, Franklin Lakes, NJ, USA) as a nitrogen source, 1.5% agar, and 0.5% Congo Red (Sigma-Aldrich) as a dye [25 (link)]. The yeast species isolated from this study were pre-cultured in YEME broth at 28℃ for 18 hr. Ten microliters of each species' cultured broth was spotted on chromogenic media. After 5 days of incubation at 28℃, the presence of a clear zone, formed on the surface of the chromogenic media due to the reaction between extracellular enzyme produced by yeast and substrate in the media was observed.
Cm cellulose
Manufactured by Merck Group
Sourced in United States
CM-cellulose is a modified form of cellulose, a naturally occurring polysaccharide. It serves as a versatile laboratory material with a range of applications in various scientific fields. The primary function of CM-cellulose is to provide a stable and inert matrix for chemical reactions, purification processes, and chromatographic techniques.
Automatically generated - may contain errors
Lab products found in correlation
Xylan, by Merck Group (2 mentions)
Deae cellulose, by Merck Group (2 mentions)
High capacity rna to cdna kit, by Thermo Fisher Scientific (2 mentions)
Polygalacturonic acid, by MP Biomedicals (1 mentions)
Starch, by Merck Group (1 mentions)
Congo red, by Merck Group (1 mentions)
D cellobiose, by Merck Group (1 mentions)
Pluronic f68, by Merck Group (1 mentions)
P cresol, by Merck Group (1 mentions)
P methoxyphenol, by Merck Group (1 mentions)
P chlorophenol, by Merck Group (1 mentions)
4 nitrophenyl acetate, by Merck Group (1 mentions)
4 o methyl d glucurono d xylan, by Merck Group (1 mentions)
Qubit double stranded dna dsdna high sensitivity assay kit, by Thermo Fisher Scientific (1 mentions)
Avicel, by Merck Group (1 mentions)
Tri reagent, by Merck Group (1 mentions)
Bradford reagent, by Merck Group (1 mentions)
Luria bertani broth, by HiMedia (1 mentions)
Sodium nitroprusside, by Thermo Fisher Scientific (1 mentions)
Sodium pentabarbitone, by Virbac (1 mentions)
11 protocols using cm cellulose
1
Extracellular Enzyme Production Screening
2
Ion Exchange Chromatography for Protease Inhibitor Purification
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The protease inhibitor fraction obtained after the dialysis of
ammonium sulphate precipitation was further purified by ion
exchange chromatography using CM cellulose (Sigma-Aldrich) as
the cation exchanger. Proteins bind to ion-exchangers due to
surface charge. These reversibly adsorbed proteins were eluted
by a step gradient of 0.1M NaCl in 0.01 M phosphate buffer with
a flow rate of 1ml/min. Peak fractions from the column were
pooled and dialyzed against 0.01M phosphate buffer (pH 7.5).
The dialyzed fractions were concentrated using amicon UF-
10KDa membrane and assayed for protease inhibitor activity,
protein content and specific activity.
ammonium sulphate precipitation was further purified by ion
exchange chromatography using CM cellulose (Sigma-Aldrich) as
the cation exchanger. Proteins bind to ion-exchangers due to
surface charge. These reversibly adsorbed proteins were eluted
by a step gradient of 0.1M NaCl in 0.01 M phosphate buffer with
a flow rate of 1ml/min. Peak fractions from the column were
pooled and dialyzed against 0.01M phosphate buffer (pH 7.5).
The dialyzed fractions were concentrated using amicon UF-
10KDa membrane and assayed for protease inhibitor activity,
protein content and specific activity.
3
Tyrosinase-Catalyzed Phenol Compound Assay
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p-Cresol, p-chlorophenol, p-methoxyphenol and other chemicals obtained from Sigma Aldrich (USA) were of the highest purity available. Mushroom Tyrosinase (50 kU, powder) was purchased from Sigma Aldrich (USA) as a solid with a specific activity of 2430 unit per mg (1 unit is defined as the enzyme activity resulting in an increase in absorbance at 280 nm of 0.001 at pH 6.5 at 25 °C in a 3 mL reaction volume containingl -tyrosine). Poly(styrene-co-divinylbenzene), glass beads, Pluronic F68, cellulose, carboxymethyl cellulose (CM-Cellulose) and triethylaminoethyl cellulose (TEAE-Cellulose) were also purchased from Sigma Aldrich (USA).
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p-Cresol, p-chlorophenol, p-methoxyphenol and other chemicals obtained from Sigma Aldrich (USA) were of the highest purity available. Mushroom Tyrosinase (50 kU, powder) was purchased from Sigma Aldrich (USA) as a solid with a specific activity of 2430 unit per mg (1 unit is defined as the enzyme activity resulting in an increase in absorbance at 280 nm of 0.001 at pH 6.5 at 25 °C in a 3 mL reaction volume containing
4
Isolation and Characterization of Paenibacillus sp. LS1
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Paenibacillus sp. LS1 was isolated from a decaying wood sample collected from Lachung village, North Sikkim district (27°40′13″N, 88°43′37″E), Sikkim, India (7 (link)). Beechwood and corncob xylan were procured from Sisco Research Laboratories Pvt. Ltd., Mumbai, India. Avicel, CM-cellulose, 4-O-methyl-d -glucurono-d -xylan, and 4-nitrophenyl acetate were procured from Sigma-Aldrich, USA. All the chemicals required for minimal medium, Luria-Bertani broth, and 3,5-dinitrosalicylic acid (DNS) reagent preparation were procured from HiMedia Laboratories, Mumbai, India, unless otherwise specified. Bradford reagent and TRI reagent were procured from Sigma-Aldrich, USA. The Qubit double-stranded DNA (dsDNA) high-sensitivity (HS) assay kit was procured from Thermo Fisher Scientific, USA.
5
Comprehensive Vascular Inflammation Assay
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The suppliers of the following compounds are as follows: Winfield red cigarettes (Phillip Morris, Australia); ebselen (Sapphire Bioscience, Australia); CM‐cellulose (Sigma‐Aldrich, USA); sodium pentabarbitone (Virbac, Australia); acridine orange/ethidium bromide (Invitrogen, USA); Kwik‐Diff® reagent 1, fixative (Thermo Fisher Scientific, USA); Hemacolour® rapid staining of blood smear (eosin solution) (Merck, USA); Hemacolour® rapid staining of blood smear (thiazine solution) (Merck, USA); RNeasy Mini Kit (Qiagen, Germany); High Capacity RNA‐to‐cDNA kit (Thermo Fisher Scientific, USA); pre‐developed TaqMan primers to: TNF‐α (ID: Mm00443258_m), IL‐6 (ID: Mm00446190_m1), NOX‐2/CYBB (ID: Mm01287743_m1), GPx‐1 (ID: Mm00656767_g1) (Thermo Fisher Scientific, USA); U46619 (Cayman Chemical, USA); ACh (Thermo Fisher, USA); sodium nitroprusside (Thermo Fisher, USA); eNOS/NOS3 RRID: AB_2533121 (Thermo Fisher Scientific, USA), 3‐NT RRID: AB110282 (Abcam, USA); Goat anti‐mouse IgG (H + L) secondary antibody, Alexa Flour Plus 488 RRID: AB_2633275 (Thermo Fisher Scientific, USA); Fluoromount‐G™, with DAPI (Thermo Fisher Scientific, USA).
6
Protein Purification Using Chromatography
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Phenylmethylsulfonylfluoride (PMSF), carboxymethyl-cellulose (CM-cellulose), diethylaminoethyl-cellulose (DEAE-cellulose), molecular weight marker kits for gel filtration and Sephacryl S-300 were purchased from Sigma Chemical Co. All other chemicals were of analytical grade.
7
Cellulose, Hematoxylin, and Nexrutine® Protocol
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CM Cellulose, hematoxylin, and eosin were purchased from Sigma (St. Louis, MO, USA), Nexrutine® (NX) was purchased from Next Pharmaceuticals (Irvine, CA). The chemicals and reagents engaged in the entire study were of the purest commercially available options.
8
Ligand Binding Assay Protocols
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All ligands used for the binding assays were purchased from Sigma-Aldrich (USA): agarose, CM-cellulose (carboxymethil cellulose), laminaritetraose (purity ≥ 90%), laminarihexaose (purity ≥ 99%), laminarin (from Laminaria digitata) and lichenan (from Cetraria islandica).
9
Screening and Assaying Cellulase and Xylanase
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Cellulase-and xylanase-producing strains were screened as previously described by Kasana et al. (2008) using LB agar containing 1% CM-cellulose (Sigma-Aldrich) or 1% xylan (xylan from beechwood, Sigma-Aldrich) for 24 h at 37∞C, respectively. For cellulase and xylanase assays, the isolates were grown overnight with agitation at 37∞C in N B supplemented with 1% CM-cellulose and 1% xylan, respectively. Enzymatic activities in culture supernatants were determined using the dinitrosalicylic acid (DNS) method (Konig et al., 2002) . One unit of activity was defined as the amount of enzyme that released 1 mmol of glucose, xylose, or their equivalents from CM-cellulose or xylan per min under assay conditions for cellulase and xylanase, respectively.
10
Psychrotolerant Marine Bacillus cereus Purification
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The psychrotolerant marine Bacillus cereus HSS, that was earlier isolated, purified and identified as a marine isolate from Mediterranean Sea, was handled in the current research [1] . Sephadex G-100 and CM-Cellulose (CMC) were obtained from Sigma (Sigma-Aldrich, USA). Fine chemicals with upper grade were applied.
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