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Example sentences for: uppsala
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The supernatant was incubated with glutathione sepharose 4B beads (Amersham Pharmacia Biotech AB, Uppsala, Sweden) and washed as previously described.
Following chromatographic separation of unincorporated 35SO 4 2-using PD-10 columns (Pharmacia, Uppsala, Sweden), the levels of radiolabeled GAGs released onto the culture media or recovered by alkaline extraction were quantitated by scintillation counting [ 24 ] .
The eggs were extracted in Celis lysis buffer (see [ 46 ] ) containing 2% (v:v) NP-40, 9.8 M urea, 100 mM dithiothreitol (DTT), 2% ampholines (pH 3.5-10), and protease inhibitors for 30 min at room temperature, according to the protocol described in [ 47 ] . Isoelectric focusing of the egg extracts (IEF) was performed using the Mini-PROTEAN II tube cell apparatus according to the manufacturer's protocol (Bio-Rad, Richmond, CA) with ampholines of pH 3.5-5 (30%), 3.5-10 (40%), 5-7 (20%), and 7-9 (10%), (Pharmacia Biotech, Uppsala, Sweden).
The van der Waals, bond, angle, dihedral, and improper dihedral parameters for all the ligands were adopted from the Hetero-compound Information Centre-Uppsala HIC-Up http://xray.bmc.uu.se/hicup [ 55 ] . The water molecule under the flaps present in all complexes was included in all steps throughout this study except for 1hvr, where the structural water was removed in the preparation of the protease to be docked with the ligand (the ligand of 1hvr, which is a urea-based inhibitor, does not bind a water molecule in this position).
Briefly, the hybridization medium contained 3 × SSC, 1.5 μg/μl poly(A) DNA (Pharmacia Biotech, Uppsala), and 0.5% SDS.
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