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Construction and expression of the eukaryotic
expressed plasmid of gene
from in IBRS-2
cells |
| JIANG Tao1, ZHANG Donglin2, NIE Hao2, ZHAO Junlong2, YAO Baoan3 |
| 1.State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University；College of Animal Science, Yangtze University；College of Veterinary Medicine, Huazhong Agricultural University; 2.State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University；College of Veterinary Medicine, Huazhong Agricultural University; 3.College of Veterinary Medicine, Huazhong Agricultural University; |
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Abstract The sequence encoding MIC3 was obtained by amplification from genomic DNA of Toxoplasma gondii RH strain and cloned into the vector pMD18-T. The target gene was subcloned into the eukaryotic vector pcDNA3.1 after the identification of pMD18-T-MIC3 by enzyme digesting, PCR amplification and sequencing. Then the target recombinant plasmids pcMIC3 were transfected into IBRS-2 cells, and the positive cells containing pcMIC3 plasmids were obtained under the selection of G418. The expressed proteins from the positive cells were detected by SDS-PAGE, Western blot and ELISA. The results showed that the DNA sequence identity was 99.9% between amplified MIC3 and that from GenBank. The molecular weight of the recombinant MIC3 protein with good immuno-activity was about 39.2 ku. These available data would lay the foundation for further studies on DNA vaccine against Toxoplasma gondii.
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Issue Date: 05 December 2008
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| 1 |
Buxton D (1998). Protozoan infections (Toxoplasmagondii, Neospora caninum and Sarcocystis spp.) in sheepand goats: recent advances. Vet Res, 29(3–4): 289–310
|
| 2 |
Càràde O, Dubremetz J F, Bout D, Lebrun M (2002). The Toxoplasma gondii proteinMIC3 requires pro-peptide cleavage and dimerization to function asadhesin. EMBO J, 21(11): 2526–2536.
doi:10.1093/emboj/21.11.2526
|
| 3 |
Choi W Y, Nam H W, Kwak N H, Huh W, Kim Y R, Kang M W, Cho S Y, Dubey J P (1997). Foodborne outbreaks of human toxoplasmosis. J Infect Dis, 175(5): 1280–1282
|
| 4 |
Dias R A, Navarro I T, Ruffolo B, Bbugni F M, Castro M V, Freire R L (2005). Toxoplasma gondii in fresh pork sausage and seroprevalence in butchers from factoriesin Londrina, Paraná State, Brazil. Rev Inst Med Trop Sao Paulo, 47(4): 185–189
|
| 5 |
Ismael A B, Sekkai D, Collin C, Bout D, Mévélec M N (2003). The MIC3gene of Toxoplasma gondii is anovel potent vaccine candidate against toxoplasmosis. Infect Immun, 71(11): 6222–6228.
doi:10.1128/IAI.71.11.6222-6228.2003
|
| 6 |
Jiang H, He S Y (2004). New advancesin the study on vaccine of Toxoplasma gondii. Foreign Med Sci Parasit Dis, 31(6): 232–265 (in Chinese)
|
| 7 |
Jiang T, Zhou Y Q, Liu Q, Nie H, Yao B A, Zhao J L (2006). Cloning and protokaryon expression of the Toxoplasma gondii MIC3 Gene. Acta Veterinaria et Zootechnica Sinica, 37(6): 587–591 (in Chinese)
|
| 8 |
Joiner K A, Dubremetz J F (1993). Toxoplasma gondii: a protozoan for the nineties. Infect Immun, 61(4): 1169–1172
|
| 9 |
Mao D G, Yang L G, Cao S X (2004). The influence factor on the efficiencyof gene immunity. Chinese Journal of Immunology, 20(5): 360–366 (in Chinese)
|
| 10 |
Mendonca A D O, Domingues P F, Silva A V D, Pezerico S B, Langoni H (2004). Detectionof Toxoplasma gondii in swine sausages. Parasitol Latinoam, 59(1–2): 42–45
|
| 11 |
Soldati D, Dubremetz J F, Lebrun M (2001). Microneme proteins: structural andfunctional requirements to promote adhesion and invasion by the apicomplexanparasite Toxoplasma gondii. Int J Parasitol, 31: 1293–1302.
doi:10.1016/S0020-7519(01)00257-0
|
| 12 |
Tenter A M, Heckeroth A R, Weiss M L (2000).Toxoplasmagondii: from animals to humans. Int J Parasitol, 30(12–13): 1217–1258.
doi:10.1016/S0020-7519(00)00124-7
|
| 13 |
Wang J P (2005). Establishment and application of the diagnostic methodsof PCR and ELISA for toxoplasmosis. Dissertationfor the Master's Degree. Wuhan: Huazhong Agricultural University, 30 (inChinese)
|
| 14 |
Yang H L, Xiao J H, Liang Y (2003). Cloning, sequencing and expressingof microneme protein 1 partial gene in Toxoplasmagondii ZS2 isolate. ChineseJournal of Preventive Medicine, 37(1): 29–32 (in Chinese)
|
| 15 |
Zhao J L (2002). Porcine parvovirus: genome sequence and cloning, expressionand gene immunity of structural protein gene. Dissertation for the Doctoral Degree. Wuhan: Huazhong Agricultural University, 40–41 (in Chinese)
|
| 16 |
Zhou J J (1999). Survey and progress of gene immunity. Foreign Medical Sciences Immunology Dis, 22(3): 119–122 (in Chinese)
|
| 17 |
Zhou Y A, Chen G J (1998). Researchprogress on protective DNA vaccine of Toxoplasmagondii. Foreign Med Sci ParasitDis, 25(5): 253–256 (in Chinese)
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