Construction of the eukaryotic coexpression vector containing Mycobacterium tuberculosis heat shock protein 70 and green fluorescent protein / 南方医科大学学报
Journal of Southern Medical University
; (12): 433-436, 2009.
Article
Dans Zh
| WPRIM
| ID: wpr-233770
Responsable en Bibliothèque :
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To construct an eukaryotic coexpression vector containing Mycobacterium tuberculosis heat shock protein 70 (mtHSP70) and enhanced green fluorescent protein (EGFP) controlled by cytomegalovirus promoter using pIRES-EGFP vector.</p><p><b>METHODS</b>The mtHSP70 gene fragment was amplified by PCR from pVAX-mtHSP70-HSV2gD using specific primers. The PCR product was cloned into the vector pMD 18-T vector, and the correct clone was selected according to DNA sequence analysis. The interested mtHSP70 gene fragment was subcloned into pCMV-IRES-EGFP vector with XhoI and EcoR I digestion. The recombinant plasmid was transfected into mouse melanoma B16 cell line, and the green fluorescent cells were detected by fluorescence microscopy and mtHSP70 expression was detected by Western blotting.</p><p><b>RESULTS</b>The recombinant plasmid obtained was confirmed by enzyme digestion. The transfected mouse melanoma B16 cells exhibited green fluorescence under fluorescence microscopy and expressed mtHSP70 protein as demonstrated by Western blotting.</p><p><b>CONCLUSION</b>The eukaryotic coexpression vector PCMV-mtHSP70-IRES-EGFP has been established to allow further investigation of the role of mtHSP70 gene in tumor immunotherapy.</p>
Texte intégral:
1
Indice:
WPRIM
Sujet Principal:
Protéines de fusion recombinantes
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Données de séquences moléculaires
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Séquence nucléotidique
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Analyse de séquence d'ADN
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Protéines du choc thermique HSP70
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Vaccins anticancéreux
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Cytomegalovirus
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Lignée cellulaire tumorale
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Protéines à fluorescence verte
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Vecteurs génétiques
Limites du sujet:
Animals
langue:
Zh
Texte intégral:
Journal of Southern Medical University
Année:
2009
Type:
Article