RESUMO
Total DNA was extracted from T. vaginalis with Chelex-100 method and used as templates for PCR. The ferredoxin gene was directionally cloned into plasmid pMD-18T vector and subcloned into eukaryotic expression vector pcDNA3. 1 (+). The transformants were screened and identified by PCR and restriction analysis. The size of amplified ferredoxin gene was 306bp and the DNA sequence of cloned gene was same with that published.
Assuntos
Células Eucarióticas/metabolismo , Ferredoxinas/genética , Proteínas de Protozoários/genética , Trichomonas vaginalis/genética , Animais , Sequência de Bases , Clonagem Molecular , Expressão Gênica , Vetores Genéticos/genética , Plasmídeos/genética , Reação em Cadeia da PolimeraseRESUMO
Trichomonas vaginalis parasitizes in human genitourinary tract. The protozoon adhering to target cell plays a critical role in its contact-dependent cytotoxicity. The enzymes synthesized by T.voginalis can hurt vaginalis epithelial cells (VECs) directly. The focal immune reaction in the location parasitized by the parasite may provide an immunologic protection. Meanwhile, inflammatory factors and immune cells may aggravate the situation. In general, the T. vaginalis-induced contact-dependent cytotoxicity is a result of the involvement of some molecular and chemical factors.
Assuntos
Apoptose , Vaginite por Trichomonas/patologia , Trichomonas vaginalis/crescimento & desenvolvimento , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Moléculas de Adesão Celular/metabolismo , Sobrevivência Celular , Cisteína Endopeptidases/metabolismo , Feminino , Humanos , Proteínas de Protozoários/metabolismo , Vaginite por Trichomonas/metabolismo , Vaginite por Trichomonas/parasitologia , Trichomonas vaginalis/metabolismoRESUMO
OBJECTIVE: To construct a recombinant plasmid containing ferredoxin gene of Trichomonas vaginalis. METHODS: Total DNA was extracted from Trichomonas vaginalis with Chelex-100 method and used as templates for PCR. Primers were designed based on the published sequence of the ferredoxin gene and used to amplify the Trichomonas vaginalis gene using PCR method. The ferredoxin gene obtained by PCR technique was directionally cloned into plasmid pMD-18T simple vector. The constructed recombinant plasmid was transferred into E. coli JM109. The transformants were screened and identified by PCR and restriction analysis. The DNA sequence of the gene was determined by Sanger's method. RESULTS: The size of amplified ferredoxin gene was 306bp. The correct recombinant plasmid was isolated and confirmed by PCR and restriction analysis. The DNA sequence of cloned gene was the same as the published sequence. CONCLUSION: The ferredoxin gene was successfully amplified and cloned into plasmid pMD-18T simple vector. The cloned ferredoxin gene could be used to produce recombinant protein and for study of its function.
