Reagents used in these experiments included T4 DNA ligase, restriction endonucleases, JM109 and BL21 E. coli strains, and goat anti-rabbit IgG alkaline phosphatase conjugate (Promega Corp., Madison, WI); a Sequenase Version 2.0 DNA Sequencing Kit (U.S. Biochemical, Cleveland, OH); [35S]dATP (> 1000 Ci/mmol; Amersham Inc., Arlington Heights, III); [a-32P]dATP (3000 Ci/mmol; Dupont/ NEN, Wilmington, DE); nitrocellulose (NitroPure) and nylon (Magna-NT) transfer membranes (Micron Separations Inc., Westboro, MA); XAR-5 and XRP film (Eastman Kodak, Rochester, NY); pGEX vector and sequencing oligonucleotides (Pharmacia Biotech, Piscataway, NJ); KlenTaq polymerase (A.B. Peptides, St. Louis, MO); P.EU. polymerase (Stratagene, La Jolla, CA); PGF2a (Lutalyse, Upjohn Co., Kalamazoo, MI); isopropyl-(3-D-thiogalactoside (IPTG; Alexis Corp., San Diego, CA); Luria Broth (Difco Laboratories, Detroit, MI); cell culture medium and fetal bovine serum (University of Missouri Cell Culture Core Facility); Tygon microbore tubing (Fisher Scientific, Pittsburgh, PA); synthetic oligonucleotides (Gibco/BRL, Gaithersburg, MD). Glutathione (GSH) agarose and all other chemicals were purchased from Sigma Chemical Co. (St. Louis, MO). buy levaquin online
Production of Recombinant ovIFN-t
Sequences encoding for the mature ovIFN-т-рЗ protein and for its truncated counterpart, from which the six carboxyl-terminal amino acids had been removed (рЗТшб), were generated through polymerase chain reaction (PCR; 94°C denaturation; 56°C annealing; 72°C synthesis; 1 min each) by using the original cDNA (p3) isolated from an ovine conceptus cDNA library (GenBank Accession Number X56341) as template, and KlenTaq and P.F.U. polymerases.