Published online Jan 14, 2005. doi: 10.3748/wjg.v11.i2.182
Revised: April 2, 2004
Accepted: May 13, 2004
Published online: January 14, 2005
AIM: To construct a recombinant murine CD40 ligand (mCD40L) eukaryotic expression vector for gene therapy and target therapy of hepatocellular carcinoma (HCC).
METHODS: mCD40L cDNA was synthesized by RT-PCR with the specific primers and directly cloned into T vector to generate middle recombinant. After digestion with restriction endonuclease, the target fragment was subcloned into the multi-clone sites of the eukaryotic vector. The constructed vector was verified by enzyme digestion and sequencing, and the product expressed was detected by RT-PCR and immunofluorescence methods.
RESULTS: The full-length mCD40L-cDNA was successfully cloned into the eukaryotic vector through electrophoresis, and mCD40L gene was integrated into the genome of infected H22 cells by RT-PCR. Murine CD40L antigen molecule was observed in the plasma of mCD40L-H22 by indirect immuno-fluorescence staining.
CONCLUSION: The recombined mCD40L eukaryotic expression vector can be expressed in H22 cell line. It provides experimental data for gene therapy and target therapy of hepatocellular carcinoma.