Cloning and expression of complementary DNA encoding an antigen of Trichinella spiralis.
Complementary DNA (cDNA) encoding a 49-kDa antigen of Trichinella spiralis was cloned, characterized, and expressed in Escherichia coli by recombinant DNA methods. As a first step, 29 residues of N-terminal amino acid sequence were determined by direct analysis of highly purified antigen. Mixed-sequence primers based on the amino acid sequence were then synthesized and used as primers to generate a partial cDNA by the polymerase chain reaction (PCR). A nearly full-length cDNA was then obtained by PCR using a specific 5'-end primer and a non-specific 3'-end primer. Complete sequence of the 1133-bp cDNA was determined. Translation of this sequence predicts an N-glycosylated polypeptide of 322 amino acids. The cDNA was expressed as a fusion protein in bacteria which could be recognized in Western blots both by sera from mice immunized with purified native 49-kDa antigen and by sera from swine infected with the parasite. These findings suggest that recombinant P49 is a potentially valuable antigen both for vaccine development and immunodiagnosis.
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Department of Parasitology, College of Veterinary Medicine, University of Georgia, Athens 30602.
Prestwood
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McGraw
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D000595:Amino Acid Sequence; D000818:Animals; D000947:Antigens, Helminth / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004247:DNA; D004277:DNA, Single-Stranded; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D006020:Glycopeptides; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000276:immunology; D013552:Swine; D014234:Trichinella / Q000235:genetics* / Q000276:immunology
D000947:Antigens, Helminth; D004277:DNA, Single-Stranded; D006020:Glycopeptides; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D004247:DNA