Human T lymphoblasts (SupT1 cells; ATCC) were grown in RPMI-1640 medium (Corning), supplemented with 10% heat-inactivated fetal bovine serum (FBS, Cellgro) and 1% penicillin/streptomycin/glutamine (Cellgro) at 37°C with 5% CO2 (g). TZM-bl reporter cells (NIH AIDS Research and Reference Reagent Program; Cat. no. 1470) and HEK293T cells were cultured in DMEM (Corning) supplemented with 10% FBS and 1% penicillin/streptomycin/glutamine at 37°C with 5% CO2 (g). Cell lines were periodically tested for mycoplasma using the MycoSensor PCR Assay Kit (Agilent).
Penicillin streptomycin glutamine (psg)
Manufactured by Corning
Sourced in United States
Penicillin/streptomycin/glutamine is a commonly used cell culture supplement that provides antibiotics and an amino acid for cell growth and maintenance. It is a sterile, liquid solution that is added to cell culture media to prevent bacterial contamination and support cell proliferation.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Corning (12 mentions)
Dulbecco modified eagle medium (dmem), by Corning (10 mentions)
Mycosensor pcr assay kit, by Agilent Technologies (4 mentions)
Rpmi 1640 medium, by Corning (3 mentions)
Rpmi 1640, by Thermo Fisher Scientific (3 mentions)
Non essential amino acids (neaa), by Corning (3 mentions)
Supt1 cells, by American Type Culture Collection (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Merck Group (2 mentions)
Cellbind plate, by Corning (2 mentions)
Penicillin streptomycin glutamine (psg), by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Omega Scientific (2 mentions)
Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Caisson (2 mentions)
Glutamax, by Thermo Fisher Scientific (2 mentions)
Basic fibroblast growth factor (bfgf), by Merck Group (1 mentions)
Insulin transferrin selenium, by Corning (1 mentions)
Dulbecco s modified eagle s medium, by Corning (1 mentions)
Penicillin streptomycin glutamax, by Thermo Fisher Scientific (1 mentions)
Hek293t cells, by Thermo Fisher Scientific (1 mentions)
28 protocols using penicillin streptomycin glutamine (psg)
1
Cell Culture Conditions for Lymphoblasts
2
Generation of A549-ACE2 Stable Cell Line
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A549 cells were obtained from the American Type Culture Collection (ATCC) and cultured in complete media containing F-12 Ham’s media (Corning), 10% fetal bovine serum (Atlanta Biologicals), 1% penicillin/streptomycin/glutamine (Corning). Calu-3 cells were also obtained from ATCC and cultured in DMEM supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin/glutamine (Corning). Both cell lines were maintained at 37°C 5% CO2. A549 cells were transduced with codon-optimized human ACE2 (Genscript) cloned into pBABE-puro [54 (link)] (Addgene). To generate the A549-ACE2 stable cell line, 107 HEK293T (ATCC) cells in T-175 flasks were transiently co-transfected with 60 μg mixture of pBABE-puro-ACE2, pUMVC, and pCMV-VSV-G at a 10:9:1 ratio using a calcium phosphate method [55 ]. Forty-eight hours post transfection, the supernatant was collected, centrifuged at 1000×g for 5 min and passed through a 0.45 μm syringe filter to remove cell debris. The filtered virus was mixed with fresh media (30% vol/vol) that included polybrene (Sigma) at a 6 μg/ml final concentration. The virus mixture was added into 6-well plates with 5×105 A549 cells/well and media was changed once more after 12 h. Transduced cells were selected in 0.5 μg/ml puromycin for 72 h, and ACE2 expression was confirmed by flow cytometry, western blot and susceptibility to HIV-1ΔEnv/SARS-CoV-2 Spike pseudovirions.
3
Generating Heterostable MDCK Cell Lines
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MDCK cells were generously provided by Prof. Jianzhu Chen (MIT), and were originally purchased from American Type Culture Collection (Manassas, VA). The identity of these cells was authenticated by STR profiling. MDCK cells were cultured at 37°C in a 5% CO2 atmosphere in DMEM (CellGro) supplemented with 10% fetal bovine serum (CellGro) and 1% penicillin/streptomycin/glutamine (CellGro). Cells were transduced with lentiviruses encoding either the DHFR.HSF1(Δ186–202) or DHFR.YFP gene. Heterostable cells expressing the construct of interest were then selected using 4 μg/mL puromycin. Single colonies were generated by diluting cells to ~40 cells per 96-well plate, expanding the resulting colonies, and functionally testing by qPCR or fluorescence microscopy in the presence or absence of trimethoprim (TMP; 10 μM). All cell lines were periodically tested for mycoplasma using the MycoSensor PCR Assay Kit from Agilent (302109).
4
Cyclic Tensile Strain on CPCs
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CPCs were seeded at a density of 4 × 105 cells/well on functionalized Bioflex plates and incubated in treatment media (Ham's F-12 (Mediatech) + 0.1 μg/mL basic fibroblast growth factor (Sigma) + 1x insulin transferrin selenium (Cellgro) + 1x penicillin-streptomycin-glutamine (Cellgro)) for 6 hours prior to the application of mechanical tension. Tensile strain was applied through a Flexcell 5000 (Flexcell International). For this, Bioflex plates were loaded onto 25 mm cylindrical loading posts. A cyclic sinusoidal strain regimen of 1 Hz and 0.5 duty cycle, with an elongation magnitude of 5, 10, or 15%, was applied to the plates for 24 hours. Cells were maintained in 5% CO2 at 37°C throughout.
5
Inducible Expression of Dominant-Negative cHSF1
6
Cell Culture Conditions for Transcriptional Activation and Nuclease Specificity Experiments
7
Temperature-Sensitive Striatal Progenitor Cells
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The creation of temperature-sensitive immortalized striatal progenitor cells from STHdhQ111/111, STHdhQ111/+ and STHdh+/+ littermate embryos, and and culture conditions, have been described (58). For the described experiment, cells were grown at the permissive temperature for the A58/U19 temperature-sensitive SV40 large-T antigen (33C) at 5% CO2. The cells were grown and maintained at 33°C in High-Glucose (25mM) DMEM (Invitrogen, cat# 11995) with 1 mM pyruvate, 10% FBS (Sigma, cat# 12306C), Penicillin-Streptomycin-Glutamine (CellGro, cat# 30-009-CI) and 400 μg/ml of G418 (Gibco, cat# 11811–031). The final concentration of glutamine in the media was 6 mM.
8
Dual Activation of Stress Pathways in HEK293 Cells
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HEK293ATF6f/XBP1s cells were cultured at 37°C in a 5% CO2 atmosphere in DMEM (CellGro) supplemented with 10% fetal bovine serum (CellGro) and 1% penicillin/streptomycin/glutamine (CellGro). Cells were generated from parental HEK293 cells (ATCC CRL-1573; authenticated by STR-profiling) as previously described (Shoulders et al., 2013 (link)). Here, 1 μM TMP and 0.1 μg/mL doxycycline were used to activate ATF6 and XBP1s, respectively. All cell lines were periodically tested for mycoplasma using the MycoSensor PCR Assay Kit from Agilent (302109).
9
Generating Stable HEK293T-REx Cell Lines
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HEK293T-REx (Invitrogen)
cells were cultured
in DMEM (CellGro) supplemented with 10% fetal bovine serum (CellGro)
and 1% penicillin/streptomycin/glutamine (CellGro). HEK293T-REx cells
were cultured in 5 mg/mL blasticidin (InvivoGen) to maintain the tet
repressor. HEK293T-REx cells were transiently transfected using calcium
phosphate. Stable, clonal HEK293T-REx cell lines were selected from
transiently transfected populations using 500 μg/mL G-418 sulfate
(Cellgro) for cells expressing pcDNA-DEST40 or pTREx-DEST30 vectors.
All stable cell lines were maintained in the appropriate selective
antibiotics.
cells were cultured
in DMEM (CellGro) supplemented with 10% fetal bovine serum (CellGro)
and 1% penicillin/streptomycin/glutamine (CellGro). HEK293T-REx cells
were cultured in 5 mg/mL blasticidin (InvivoGen) to maintain the tet
repressor. HEK293T-REx cells were transiently transfected using calcium
phosphate. Stable, clonal HEK293T-REx cell lines were selected from
transiently transfected populations using 500 μg/mL G-418 sulfate
(Cellgro) for cells expressing pcDNA-DEST40 or pTREx-DEST30 vectors.
All stable cell lines were maintained in the appropriate selective
antibiotics.
10
Quantifying Viral Titers in Cells
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Viral titers in infectious inoculum and in indicated organs were determined as described previously65 (link). In brief, homogenized samples were serially diluted (10-fold) onto MC57G cells seeded into 24-well plates (105/well) in MEM supplemented with 5% FCS and 1% PSG (Corning and Gibco). Infected cells were overlayed 4 h later with a 1:1 mix of 2% methylcellulose (Sigma) and 5% FCS/DMEM medium (Gibco), and incubated for an additional 48 h at 37 °C. Cells were fixed with 4% PFA, permeabilized with 1% Triton X-100, and stained with rat anti-LCMV VL-4 and peroxidase-conjugated goat anti-rat IgG (Jackson ImmunoResearch). For visualization OPD (Sigma) substrate was applied for >30 min at RT.
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