CELL LINES
Contributor Information
- Name Jose Luis Pedraz ; Laura Saenz del Burgo ; JesÄ?s Ciriza
- Institute University of the Basque Country (EHU)
Tool Details
- Tool name: GFP-expressing D1-MSC cell line
- Tool type: Cell Lines
- Parental cell line: D1-MSC
- Organism: Mouse
- Tissue: Bone Marrow
- Growth properties: Adherent cells
- Model: Stem Cells
- Conditional: No
- Description: Mesenchymal stem cells (MSCs) are multipotent stem cells found in bone marrow that are important for making and repairing skeletal tissues, such as cartilage, bone and the fat found in bone marrow. D1-MSC's are a cell line of murine MSCs from bone marrow. D1-MSCs were transduced with the lentiviral construct pSIN-EF2-GFP-Pur, which codifies GFP from Aequorea coerulescens under the human elongation factor-1 alpha (EF-1 alpha) promoter, linked to puromycin resistance through an internal ribosome entry site (IRES) motif.
- Research area: Developmental biology; Cell biology
- Production details: Complementary DNA (cDNA) of GFP was cloned from the plasmid pAcGFP1-N1 containing GFP from Aequorea coerulescens and cloned on a lentiviral vector (pSIN) to give pSIN-EF2-GFP-Pur. This was used to transfect HEK293 cells from which the supernatant-containing lentivirus was harvested after 3 days. This supernatant was used to transduce D1-MSCs for 1 month after which GFP expression was confirmed by fluorescence microscopy. Lack of virus production by the now GFP-expressing D1-MSCs was confirmed through attempts at using their supernatant to infect other cells. Insertion of GFP cDNA sequence into the genome was determined by PCR.
- For Research Use Only
Target Details
- Target: GFP
Application Details
Handling
- Format: Frozen
- Growth medium: DulbeccoÄÂ?´s Modified EagleÄÂ?´s Medium (DMEM) high glucose media supplemented with 10% heat-non-inactivated fetal calf serum and 1 unit/mL of penicillin-streptomycin and puromycin at 12.5 ÄÂÄ?g/mL.
- Shipping conditions: Dry ice
Documentation
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