Abstract
To study subcellular localization of α18- and α12-giardin in Giardia lamblia trophozoites, the α18- and α12-giardin genes were amplified from G. lamblia assemblage A, respectively. The PCR products were cloned into the prokaryotic expression vector pET-28a(+), and the positive recombinant plasmids were transformed into E. coli Rosetta (DE3) strain for the expression, and expressed α18- and α12-giardin fusion protein were purified by Ni-Agarose resin, respectively. Mice were immunized with purified fusion proteins for preparation of polyclonal antibody, and then the subcellular localization of α18- and α12-giardin was determined by fluorescence immunoassay. Results showed that the concentrations of purified α18- and α12-giardin fusion proteins were 1.20 and 0.86 mg/ml, respectively. The titers of anti-α18- and anti-α12-giardin polyclonal antibody were both as high as 1:25600 dilutions. Immunofluorescent analysis showed that α18- and α12-giardin proteins were mainly localized at four pairs of flagella and the cytoplasm of G. lamblia trophozoites, suggesting that α18- and α12-giardin are the flagella and cytoplasm-associated proteins, respectively. The above information would lay the foundation for research about the crystal structure and biological function of α18- and α12-giardin.
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Acknowledgments
This work was supported by a grant from the National Natural Science Foundation of China (grant no. 31272551). The authors would like to thank Prof. Zhaorong Lun from Southern China Research Center of Parasitic Biology, Sun Yat-Sen University, China, for offering the G. lamblia trophozoites assemblages A.
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Wu, S., Pan, W., Shi, X. et al. Immunolocalization of α18- and α12-giardin in Giardia lamblia trophozoites. Parasitol Res 115, 4183–4187 (2016). https://doi.org/10.1007/s00436-016-5194-z
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DOI: https://doi.org/10.1007/s00436-016-5194-z