Vero, HEK-293T, and BHK-21 cells were obtained from the American Type Culture Collection and propagated in DMEM supplemented with 5% (Vero and BHK-21) or 10% (HEK-293T) FBS (Omega Scientific), 100 U/ml penicillin, 100 μg/ml streptomycin and 10 mM HEPES. All cell lines were tested and judged free of mycoplasma contamination using a commercial kit. The plasmids pKR780–2-EEEV, pKR780–2-VEEV, and pKR780–2-WEEV are comprised of the codon-optimized pE2–6K-E1 genes of EEEV FL93–939, VEEV TrD, and WEEV CB87, respectively, under the control of a chicken β-actin promoter, which have been cloned into the pCAGGS expression vector (Addgene). Replication-competent SINV chimeric viruses were constructed by replacing the SINV TR339 structural protein genes with EEEV FL93–939 structural protein genes under control of the SINV subgenomic promoter in the TR339 cDNA clone47 (link). The cDNA clones of EEEV TrD FL93–939 WT and nanoluciferase-expressing challenge viruses have been described30 (link),48 (link).
Bhk 21
Manufactured by Omega Scientific
The BHK-21 is a cell line derived from baby hamster kidney cells. It is commonly used in cell culture applications to support the growth and maintenance of various cell types.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Omega Scientific (3 mentions)
Hek 293t, by Omega Scientific (3 mentions)
Hek293t, by American Type Culture Collection (3 mentions)
Bhk 21, by American Type Culture Collection (3 mentions)
Pcaggs expression vector, by Addgene (2 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Iscove s modified dulbecco s medium, by Thermo Fisher Scientific (1 mentions)
Hepes, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle medium, by Thermo Fisher Scientific (1 mentions)
3 protocols using bhk 21
1
Plasmid Construction and Virus Generation
2
Cell Line Culture Protocols
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Cell lines were maintained at 37°C in the presence of 5% CO2. Vero (American Type Culture Collection [ATCC] CCL-81), HEK293T (ATCC CRL-3216), BHK-21 (ATCC CCL-10), or ΔB4galt7 N2a (Ma et al., 2020 (link)) cells were passaged in growth medium Dulbecco's Modified Eagle Medium (DMEM [Invitrogen] supplemented with 5% [Vero and BHK-21] or 10% [HEK293T and ΔB4galt7 N2a] heat-inactivated FBS [Omega Scientific], 100 U/ml penicillin, 100 U/ml streptomycin, and 10 mM Hepes [Invitrogen]). Gibco Expi293F (Thermo Fisher Scientific) cells were maintained at 37°C in Expi293 Expression Medium. Expi293F cells were authenticated by the ATCC cell line authentication service using short tandem repeat analysis. Hybridoma cell lines were grown in Iscove's Modified Dulbecco's Medium (Thermo Fisher Scientific) supplemented with 20% FBS (Hyclone), 1 mM sodium pyruvate, 100 U/ml penicillin, and 100 U/ml streptomycin (Invitrogen). HMMA 2.5, B95.8, and ExpiCHO cell lines were maintained as previously described (Williamson et al., 2020 (link), 2021 (link)). All cell lines were confirmed as mycoplasma negative using Washington University Induced Pluripotent Stem Cell Core Facility services or using a universal mycoplasma detection kit (ATCC 30-1012K).
3
Plasmid Construction and Virus Generation
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