Cloning and analysis of 20K cyclophilin gene of Cryptosporidium parvum

Objective To acquire 20K cyclophihin(CyP) gene of Cryptosporidium parvum(C.parvum) and to analyze the nucleotide and amino acid sequences of the gene.Methods We constructed animal models of NJ strain of C. parvum infection with Kunming mice.We designed two pairs of primers based on the known gene sequences of 20K CyP of C. parvum Iowa Ⅱ strain and amplified it from the C. parvum NJ strain by nested PCR method,then cloned it into the pMD18-T vectors.The positive recombinant plasmid pMD18-T-20K CyP was acquired after the identification by PCR and double enzyme digestion methods.Results We achieved right recombinant plasmid pMD18-T-20K CyP by the identification of PCR and double enzyme digestion method.The sequencing results of the nucleotides showed that the 20K CyP gene of the C.parvum NJ strain was 519 bp in full length,encoding 173 amino acids.The gene had been logined to GenBank database with the accession number of JQ284431.And the sequence analyses indicated that it had 100% homology in amino acids with the Iowa II strain.After analyzing its protein structure domain,there were specific cyclophilin A,B,Hlike PPIase domain which were similar to human cyclophilins A,B,and H.Conclusion The 20K CyP gene of C.pravum NJ strain was successfully cloned,and the structure of 20K CyP is highly conserved in C. parvum.