Development of a novel retrovirus vector system for hematopoietic Stem cells

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The aims of the project was as follows ;1. Evaluation of the gene expression by a novel retroviral vector which consisted of the SFFVp LTR and the MESV leader sequences.2. Establishment and the evaluation of novel packaging cells based on stromal cells.3. Evaluation of the pseudo-typed retroviral vectors based on SFFVp with VSV G-protein.We have obtained the following results ;1. A series of retroviral vectors were constructed. Using the packaging cells based on NIH/3T3, these vector-producing cells were isolated. Expression levels of these vectors were compared in hematopoietic progenitors. The highest expression of a marker gene among these was obtained by the FMEV-type vector, which contained the SFFVp LTR and the MESV leader sequences.2. Plasmid DNAs of the expression vector carrying gal/pol sequences and that carrying the MoMLV env sequences were introduced into a stromal cell line, MS-5. These novel stromal cell-based packaging cells were found to be unstable to produce high titer of viral vectors.3. SFFVp based retroviral vectors could be pseudo-typed with VSV G-protein and these particles successfully transduced the cells.