The U. diversum ATCC 49782 strain was isolated from a cow with granular vulvovaginitis in Ontario, Canada [39 (link)], and 45 U. diversum isolates (Table S1 ) were provided by the Mycoplasma Laboratory of the Institute of Biomedical Sciences, University of São Paulo, Brazil (USP). Some strains were isolated from cows with granulomatous vulvovaginitis, whereas others were isolated from the semen of healthy bulls. The isolates were obtained from the following four states: 19 from São Paulo, two from Mato Grosso do Sul, one from Minas Gerais, and 22 from Bahia. Next, 1 mL of each sample, previously stored in Ureaplasma medium (UB) containing 21 g of Difco PPLO broth, 2 mL of 1% phenol red dye, 20 mL of unheated horse serum ((Invitrogen®, Waltham, MA, USA), 10 mL of 25% fresh yeast extract, 1 mL of 10% urea solution (Sigma®, St. Louis, MO, USA), and 200,000 units/mL of penicillin G (Sigma®) per litre of the medium, was grown in 9 mL of the same medium at 37 °C for 24–48 h [6 (link),29 (link)]. Bacterial DNA was extracted using a NucleoSpin kit (Macherey-Nagel, Düren, Germany), according to the manufacturer’s instructions.
Urea solution
Manufactured by Merck Group
Sourced in United States
Urea solution is a laboratory reagent that is commonly used in various analytical and diagnostic applications. It is an aqueous solution of the organic compound urea, which is the main nitrogenous waste product in the urine of mammals. The urea solution is typically used as a standard or a control material in procedures that measure urea concentration, such as those employed in clinical chemistry and biochemistry.
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Lab products found in correlation
Penicillin g, by Merck Group (1 mentions)
Nucleospin kit, by Macherey-Nagel (1 mentions)
α1 6 mannosidase, by New England Biolabs (1 mentions)
O glycosidase, by New England Biolabs (1 mentions)
α2 3 6 8 neuraminidase, by New England Biolabs (1 mentions)
Mrcf0r030, by Merck Group (1 mentions)
α1 2 fucosidase, by New England Biolabs (1 mentions)
Ammonium bicarbonate, by Merck Group (1 mentions)
Pngase f, by New England Biolabs (1 mentions)
2 protocols using urea solution
1
Isolation and Characterization of Ureaplasma diversum from Bovine Samples
2
Glycan Modification Analysis in BMDCs
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The 500 µg of total proteins extracted from wild-type or wls-null (wlsfx/fx) BMDC were added to a 30 k centrifugal filter unit (Merck Millipore, MRCF0R030). Then, urea solution (8 M; Sigma-Aldrich, U5378) was added so as to bring the total volume to 400 µl. Samples were centrifuged at 16,000 x g for 15 min to remove cytosolic contaminants, and 200 µl of urea solution was added to wash it twice. Next, after adding 200 µl of ammonium bicarbonate (50 mM; Sigma-Aldrich, 09830), the tubes were centrifuged at 16,000 x g for 15 min to exchange buffer. The different glycan modifications were released from the samples by the addition of 6 UN of PNGase F (P0704S), O-glycosidase (P0733S), α1-6 mannosidase (P0727S), α2-3,6,8 neuraminidase (P0720S), or α1-2 fucosidase (P0724S), all from New England Biolabs, in 200 μl of NH4HCO3 solution (25 mM) at 37°C; they were digested overnight (16 h) under shaking conditions and finally centrifuged at 16,000 x g for 15 min to get the released glycans in a new tube.
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