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| 产品名称： | ProPak-A.52 Clone #52 PP-A.52 |
|---|---|
| 商品货号： | TS211088 |
| Organism： | Homo sapiens, human |
| Cell Type： | transformed with adenovirus 5 DNA |
| Product Format： | frozen |
| Culture Properties： | adherent |
| Biosafety Level： | 2 Cells contain adenovirus and CMV viral sequences
Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country. |
| Disease： | Leukemia |
| Age： | fetus |
| Applications： | The ProPak packaging cell lines produce either murine leukemia virus (MLV) xenotropic particles (ProPak-X cells; ATCC CRL-12007) or amphotropic particles (ProPak-A cells; ATCC CRL-12006 and TS211088). They were derived from the human embryonic kidney line, 293 (see ATCC CRL-1573). ProPak-based producer cells were demonstrated to be free of replication-competent retrovirus (RCR) by stringent testing. Consistently higher transduction of target cells was achieved with ProPak-derived amphotropic vector than with PA317-packaged amphotropic vector. |
| Storage Conditions： | liquid nitrogen vapor phase |
| Disclosure： | This material is cited in a US or other Patent and may not be used to infringe the claims. Depending on the wishes of the Depositor, ATCC may be required to inform the Patent Depositor of the party to which the material was furnished. This material may not have been produced or characterized by ATCC. |
| Derivation： | They were derived from the human embryonic kidney line, 293 (see ATCC CRL-1573). The CMV promoter was excised (EcoRI/XhoI, blunt-ended), and replaced with the MoMLV LTR (Asp 718/HindIII, blunt-ended) from plasmid pVH2. The amphotropic envelope-encoding plasmid, pCMVEa, was transfected into ProGag cells and clones yielding high transduction efficiencies were isolated. One of these, clone number 52 (ProPak-A.52), was deposited as CRL-12479. Consistently higher transduction of target cells was achieved with ProPak-derived amphotropic vector than with PA317-packaged amphotropic vector. |
| Clinical Data： | To construct the ProPak-X and the ProPak-A-52 cell lines, the ATG in the splice donor/splice acceptor of pCMV plasmid was mutated to ACG. |
| Comments： | The ProPak packaging cell lines produce either murine leukemia virus (MLV) xenotropic particles (ProPak-X cells; ATCC CRL-12007) or amphotropic particles (ProPak-A cells; ATCC CRL-12006 and TS211088). They were derived from the human embryonic kidney line, 293 (see ATCC CRL-1573). To construct the ProPak-X and the ProPak-A-52 cell lines, the ATG in the splice donor/splice acceptor of pCMV plasmid was mutated to ACG. The CMV promoter was excised (EcoRI/XhoI, blunt-ended), and replaced with the MoMLV LTR (Asp 718/HindIII, blunt-ended) from plasmid pVH2. The beta-galactosidase gene was replaced by the gag-pol ORF (NotI fragment) to generate pMoMLVgp. pMoMLVgp was co-transfected with pHA58 into 293 cells by calcium phosphate co-precipitation and hygromycin B-resistant cells were selected. Clones were screened for the level of Gag secretion and one clone secreting high levels of Gag was selected (designated ProGag); this clone yielded high viral titers in transient transfection. The amphotropic envelope-encoding plasmid, pCMVEa, was transfected into ProGag cells and clones yielding high transduction efficiencies were isolated. One of these, clone number 52 (ProPak-A.52), was deposited as CRL-12479. ProPak-based producer cells were demonstrated to be free of replication-competent retrovirus (RCR) by stringent testing. The highest transduction of human hematopoietic progenitor cells was achieved with vector supernatant generated from a coculture of the ProPak-X and ProPak-A cell lines. Consistently higher transduction of target cells was achieved with ProPak-derived amphotropic vector than with PA317-packaged amphotropic vector. |
| Complete Growth Medium： | The base medium for this cell line is ATCC-formulated Dulbeccos Modified Eagles Medium, Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10%.
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| Subculturing： | Protocol: Remove medium. Do not rinse. Cells detach easily. Add 2.0 to 3.0 ml of 0.25% trypsin, 0.53 mM EDTA solution. Allow the flask to sit at room temperature (or at 37C) until the cells detach. Centrifuge the cell suspension at 1000 rpm for 10 minutes, resuspend the pellet in fresh medium, aspirate and dispense into new flasks. Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:10 is recommended Medium Renewal: Every 2 to 3 days |
| Cryopreservation： | Freeze medium: Complete growth medium 95%; DMSO, 5% Storage temperature: liquid nitrogen vapor phase |
| Culture Conditions： | Temperature: 37.0°C |
| STR Profile： | Amelogenin: X CSF1PO: 11,12 D13S317: 12,14 D16S539: 9 D5S818: 8,9 D7S820: 11,12 THO1: 7,9.3 TPOX: 11 vWA: 16,19 |
| Name of Depositor： | SyStemix, Inc. |
| Deposited As： | human |
| U.S. Patent Number： | |
| References： | Rigg RJ, et al. Method for obtaining retroviral packaging cell lines producing high transducing efficiency retroviral supernatant. US Patent 6,017,761 dated Jan 25 2000 Forestell SP, et al. Novel retroviral packaging cell lines: complementary tropisms and improved vector production for efficient gene transfer. Gene Ther. 4: 600-610, 1997. PubMed: 9231077 |