Isolation and characterization of cDNA clones encoding a 32-kDa dense-granule antigen of Sarcocystis muris (Apicomplexa)

Abstract

A monoclonal antibody (2F4) directed against a 32-kDa dense-granule antigen of Sarcocystis muris cyst merozoites (bradyzoites) was used to screen a lambda ZAP cDNA expression library. A clone with an insert of 1.4 kb in length (DG 32/1) was isolated. A fusion protein derived from bacteria harbouring the recombinant plasmid DG 32/1 reacted with monoclonal antibody (mAb) 2F4. Southern blot hybridization suggests that the gene is present as a single copy. On Northern blots, a single mRNA species of 1.8 kb was detected by a cDNA-derived probe. In addition, we isolated a full-length clone (DG 32/PH1) by screening the cDNA library with a non-radioactive-labelled cDNA probe. The nucleotide sequence of DG 32/PH1 comprises 1.57 kb. It contains an open reading frame of 882 bp with a coding capacity of approximately 32 kDa. The hypothetical polypeptide consists of a putative N-terminal signal peptide and the mature protein sequence. The occurrence of an N-terminal signal sequence is consistent with the observation that the 32-kDa protein of S.␣muris is secreted from the dense granules.