Cloning, Nucleotide Sequencing and Bioinformatics Study of NcSRS2 Gene, an Immunogen from Iranian Isolate of Neospora caninum
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Abstract
Background: Neosporosis is caused by an obligate intracellular parasitic protozoa Neospora caninum which infect variety of hosts. NcSRS2 is an immuno-dominant antigen of N. caninum which is consi-dered as one of the most promising targets for a recombinant or DNA vaccine against neosporosis. As no study has been carried out to identify the molecular structure of N. caninum in Iran, as first step, we prepared a scheme to identify this gene in this parasite in Iran.
Methods: Tachyzoite total RNA was extracted and cDNA was synthesized and NcSRS2 gene was amplified using cDNA as template. Then the PCR product was cloned into pTZ57R/T vector and transformed into E. coli (DH5α strain). Finally, the recombinant plasmid was extracted from trans-formed E. coli and sequenced. Bioinformatics analysis also carried out.
Results: The PCR product of NcSRS2 gene was sequenced and recorded in GenBank. The deduced amino acid sequence of NcSRS2 in current study was compared with other N. caninum NcSRS2 and showed some identities and differences.
Conclusion: NcSRS2 gene of N. caninum successfully cloned in pTZ57R/T. Recombinant plasmid was confirmed by sequencing, colony PCR and enzymatic digestion. It is ready to express recombi-nant protein for further studies.
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