Flp-In-3T3 BioVector NTCC质粒载体菌种细胞基因保藏中心

Flp-In-3T3

BioVector NTCC质粒载体菌种细胞蛋白抗体基因保藏中心www.biovector.net

Description of Flp-In™ cell linesAll of the Flp-In™ cell lines (except Flp-In™-CHO; see the following section) contain a single integrated FRT site and stably express the lacZ-Zeocin™ fusion gene from the pFRT/lacZeo plasmid under the control of the SV40 early promoter (see the following diagram). The location of the FRT site in each Flp-In™ cell line has not been mapped, but is presumed to have integrated into a transcriptionally active genomic locus as determined by generation of a Flp-In™ expression cell line containing the pcDNA™5/FRT/CAT or pEF5/FRT/GW-CAT control plasmid. The Flp-In™ cell lines should be maintained in medium containing Zeocin™ Selection Antibiotic (see Media for cell lines, page 7).For more information about pFRT/lacZeo, refer to the Flp-In™ System manual. For information about pcDNA™5/FRT/CAT or pEF5/FRT/GW-CAT, refer to the pcDNA™5/FRT or pEF5/FRT/V5-DEST™ manuals, respectively.  The Flp-In™-CHO cell line contains a single integrated FRT site and stably expresses the lacZ-Zeocin™ fusion gene from the pFRT/lacZeo2 plasmid. Note that pFRT/lacZeo2 contains a mutated SV40 early promoter (PSV40Δ) which is severely abrogated in its activity. The SV40Δ early promoter in pFRT/lacZeo2 exhibits approximately 60-fold less activity than the wild-type SV40 early promoter in pFRT/lacZeo. Because of the minimal activity of the SV40Δ promoter, we expect that stable transfectants expressing the lacZ-Zeocin™ gene from pFRT/lacZeo2 should contain FRT sites which have integrated into the most transcriptionally active genomic loci. The location of the FRT site in the Flp-In™-CHO cell line has not been mapped, but has been demonstrated to have integrated into a highly transcriptionally active genomic locus as determined by generation of a Flp-In™ expression cell line containing the pcDNA™5/FRT/luc (luciferase-expressing) control plasmid. The Flp-In™-CHO cell line should be maintained in medium containing Zeocin™ Selection Antibiotic (see Media for cell lines, page 7). For more information about pFRT/lacZeo2 and the SV40Δ early promoter, refer to the pFRT/lacZeo2 manual.    Important guidelines•FBS does not need to be heat inactivated for use with these cell lines.•Cell lines should be maintained in medium containing Zeocin™ Selection Antibiotic at the concentrations listed in the previous section.•If adherent cells (e.g., Flp-In™-293, Flp-In™-CV-1, Flp-In™-CHO, Flp-In™-3T3, Flp-In™-BHK) are split at a 1:5 to 1:10 dilution, they will generally reach 80–90% confluence in 3–4 days.•Suspension Flp-In™-Jurkat cells will demonstrate optimal growth characteristics if maintained at a cell density between 1 × 105 cells/mL and 1 × 106 cells/mL.•When maintaining Flp-In™-Jurkat cells in suspension culture, do not allow the medium to turn yellow; this indicates that cells have reached too high a density or that the medium is depleted of nutrients. If this occurs, either add fresh complete media to the cells or passage them. BioVector NTCC质粒载体菌种细胞蛋白抗体基因保藏中心www.biovector.net    Flp-In™-293D-MEM (high glucose) 10% FBS* 2 mM L-glutamine 1% Pen-Strep (optional)100 μg/mL Zeocin™ Selection Antibiotic90% complete medium10% DMSOFlp-In™-CV-1D-MEM (high glucose) 10% FBS* 2 mM L-glutamine 1% Pen-Strep (optional)100 μg/mL Zeocin™ Selection Antibiotic90% complete medium10% DMSOFlp-In™-CHOHam’s F12 10% FBS* 2 mM L-glutamine 1% Pen-Strep (optional)100 μg/mL Zeocin™ Selection Antibiotic90% complete medium10% DMSOFlp-In™-BHKD-MEM (high glucose) 10% FBS* 2 mM L-glutamine 1% Pen-Strep (optional)100 μg/mL Zeocin™ Selection Antibiotic90% complete medium10% DMSOFlp-In™-3T3D-MEM (high glucose) 10% donor calf serum 2 mM L-glutamine 1% Pen-Strep (optional)100 μg/mL Zeocin™ Selection Antibiotic90% complete medium10% DMSOFlp-In™-JurkatRPMI 1640 10% FBS* 2 mM L-glutamine 1% Pen-Strep (optional)100 μg/mL Zeocin™ Selection Antibiotic90% complete medium10% DMSO      [Supplier来源]  http://www.biovector.net