Molecular cloning and expression of the full-length tropomyosin gene from Trichinella spiralis.
A clone, designated as TsTM, was selected from the cDNA library of newborn larvae (NBL) of Trichinella spiralis through immunoscreening against infected sera. The clone contained a cDNA transcript of 855 bp in length with a single open reading frame, which encoded 285-amino acids (33 kDa in the estimated molecular weight). A sequence analysis revealed that the clone TsTM encoded the full-length of tropomyosin gene. The phylogenetic analysis of the tropomyosin gene was in good agreement with the classical taxonomical position of T. spiralis. The fusion proteins encoded by the clone TsTM were produced in an Escherichia coli expression system and affinity purified, and the antibody was raised against the protein for the following studies. The antibody against the fusion protein positively bound to the hypodermal muscle layer in immunolocalization analysis, and the 35 kDa band in crude extracts of muscle larvae but not in excretory and secretory (ES) products on Western blots. The antigenicity of the clone TsTM was recognized by host mice but exhibited little species specificity.
Nakada
T
T
Department of Parasitology, Gifu University School of Medicine, Tsukasa 40, 500-8705, Japan.
Nagano
I
I
Wu
Z
Z
Takahashi
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D000909:Antibodies, Helminth / Q000032:analysis; D015153:Blotting, Western / Q000379:methods; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D007150:Immunohistochemistry; D007814:Larva; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009348:Nematoda / Q000235:genetics; D010802:Phylogeny; D016415:Sequence Alignment; D017160:Trichinella spiralis / Q000235:genetics* / Q000276:immunology; D014335:Tropomyosin / Q000235:genetics* / Q000276:immunology
D000909:Antibodies, Helminth; D014335:Tropomyosin