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Phenotype and T Cell Stimulating Activity of Dendritic Cells Expanded from Human Bone Marrow and Cord Blood CD34~( ) Hematopoietic Progenitor Cells
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Abstract:
Objective: To amplify the tk gene of HSV- I strain Stocker and clone into a eukaryotic expressing plasmid. A high effective expressing vector containing HSV-I tk gene would be constructed. Methods: PCR amplification was performed using primers based on tk gene sequence of HSV-I strain CL101, nucleotide of strain Stocker as template. PCR product was cloned into plasmid pUCl 19 and the sequence of tk gene was analyzed. The tk gene was then cloned into a high effective eukaryotic expressing plasmid which contains CMV immediate early gene promoter. The recombinant pCR3-tk was further identified by restriction digest. Results: A 1427 bp DNA fragment was amplified. Sequence analysis and restriction digest demonstrated that the fragment cloned contained complete HSV-I tk and indicated a 98.5% identity between the cloned tk gene and that from HSV-I CL101. pCR3-tk vector was identified by restrition digest. Conclusion: The identity between the cloned tk gene and that from HSV-I CL101 is very high. The recombinant of pCR3-tk is a high effective expressing eukaryotic vector. It will play a fundamental role in further study of tumor gene therapy with HSV- I tk/GCV system expecially mediated by non-viral vectors such as cationic liposomes.
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Clc Number:
R730.5
