Lentiviral constructs expressing human SMN or GFP from the CMV promoter were generated by standard cloning techniques using the pRRLSIN.cPPT.PGK-GFP.WPRE vector (Addgene plasmid 12252) as initial backbone. Viral stocks pseudotyped with the vesicular stomatitis G protein (VSV-G) were prepared by transient co-transfection of HEK293T cells (Open Biosystems) using the ViraPower™ Lentiviral Packaging Mix (Invitrogen) following manufacturer’s instructions. Supernatant was collected 48 hours post transfection. Lentivirus was concentrated by ultracentrifugation at 19,500 rpm for 2.5 hours at 20°C, reconstituted in PBS, and stored in aliquots at −80°C. The viral titer was determined with the Lenti-XTM qRT-PCR Titration Kit (Clontech). A multiplicity of infection of 1:1 was used to transduce HeLa cells grown in DMEM with high glucose (Invitrogen) containing 10% of FBS (HyClone), 2 mM glutamine (Gibco), and 1% penicillin and streptmomycin (Gibco).
Penicillin and streptmomycin
Manufactured by Thermo Fisher Scientific
Penicillin and streptmomycin are antibiotics commonly used in microbiology and molecular biology laboratories. They function as antimicrobial agents that inhibit the growth of bacteria.
Automatically generated - may contain errors
Lab products found in correlation
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Glutamine, by Thermo Fisher Scientific (2 mentions)
High glucose, by Thermo Fisher Scientific (2 mentions)
Virapower lentiviral packaging mix, by Thermo Fisher Scientific (2 mentions)
Lenti xtm qrt pcr titration kit, by Takara Bio (2 mentions)
2 protocols using penicillin and streptmomycin
1
Lentiviral Transduction of HeLa Cells
2
Lentiviral Transduction of HeLa Cells
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Lentiviral constructs expressing human SMN or GFP from the CMV promoter were generated by standard cloning techniques using the pRRLSIN.cPPT.PGK-GFP.WPRE vector (Addgene plasmid 12252) as initial backbone. Viral stocks pseudotyped with the vesicular stomatitis G protein (VSV-G) were prepared by transient co-transfection of HEK293T cells (Open Biosystems) using the ViraPower™ Lentiviral Packaging Mix (Invitrogen) following manufacturer’s instructions. Supernatant was collected 48 hours post transfection. Lentivirus was concentrated by ultracentrifugation at 19,500 rpm for 2.5 hours at 20°C, reconstituted in PBS, and stored in aliquots at −80°C. The viral titer was determined with the Lenti-XTM qRT-PCR Titration Kit (Clontech). A multiplicity of infection of 1:1 was used to transduce HeLa cells grown in DMEM with high glucose (Invitrogen) containing 10% of FBS (HyClone), 2 mM glutamine (Gibco), and 1% penicillin and streptmomycin (Gibco).
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