X tremegene hp transfection reagent

Human neuroblastoma SH-SY5Y cells (ATCC, Manassas, VA) were grown in DMEM containing 10% FBS (vol/vol) and antibiotics (penicillin-streptomycin 100 U/ml, ThermoFisher Scientific). Cells were propagated in a humidified atmosphere consisting of 5% CO2/95% air and maintained at 37 °C. For transient transfections of the indicated vectors, X-tremeGENE HP transfection reagents (Roche) were used according to the manufacturer’s instructions. Primary cortical cell cultures were prepared from gestational day 15 mouse embryos as previously described [6 (link)]. Experiments were performed at DIV (day in vitro) 14.

Human neuroblastoma SH-SY5Y cells (ATCC, Manassas, VA, USA) were grown in complete media containing DMEMs, 10% fetal bovine serum (vol/vol), and antibiotics (penicillin-streptomycin 100 U/mL, Thermo Fisher Scientific, Waltham, MA, USA). The atmospheric condition used to preserve the cells consisted of 5% CO₂/95% air at 37 °C and maintained a humidified environment. X-tremeGENE HP transfection reagents (Roche, Basel, Switzerland) were used for transient transfection of DNA constructs (pCMV-tTA, TetP-β23, and pTet-dual2 [5 (link)]) following the manufacturer’s instructions.

Human embryonic kidney cells (HEK-293T) and human neuroblastoma SH-SY5Y cells (ATCC, Manassas, VA) were grown in DMEM containing 10% (vol/vol) fetal bovine serum (FBS) and antibiotics. Cells were cultured at 37 °C in a humidified incubator supplied with 5% CO₂/95% air. For transient transfection, cells were transfected with indicated constructs using X-tremeGENE HP transfection reagents (Roche) according to the manufacturer’s instructions. Unless otherwise indicated, lysates were prepared at 48 h post transfection.

Jurkat cells were transfected with the pLTR-luc and the pLTR-NFκBmut-luc luciferase reporter episomal vectors using JetPEI TM (POLYplus) method according to the manufacturer’s protocol. Twenty-four hours post-transfection, cells were mock-treated or treated with the different compounds as indicated. At 24 hours post-treatment, cells were lysed and assayed for luciferase activity (Promega) as previously described [66 (link)]. Luciferase activities were normalized with respect to protein concentrations.
HeLa cells were transiently transfected with the Hex1(-104)Luc reporter plasmid [48 (link)] by X-tremeGENE HP Transfection Reagents (Roche). At 24 hours post-transfection, cells were mock-treated or treated with the different compounds as indicated. At 24 hours post-treatment, cells were lysed and assayed for luciferase activity (Promega). Luciferase activities were normalized with respect to protein concentrations.

Human neuroblastoma SH-SY5Y cells (ATCC, Manassas, VA) were grown in DMEM containing 10% FBS (vol/vol) and antibiotics (penicillin-streptomycin 100 U/mL, ThermoFisher Scientific, Waltham, MA, USA). Cells were propagated in a humidified atmosphere consisting of 5% CO₂/95% air and maintained at 37 °C. For transient transfections of the indicated vectors, X-tremeGENE HP transfection reagents (Roche, Mannheim, Germany) were used according to the manufacturer’s instructions.

Human neuroblastoma SH-SY5Y cells (ATCC) and human embryonic kidney (HEK)-293T cells were grown in Dulbecco’s Modified Eagle’s Medium (DMEM) containing 10% (v/v) fetal bovine serum (FBS) and antibiotics. Cells were cultured at 37 °C in a humidified incubator supplied with 5% CO₂/95% air. For transient transfection, cells were transfected with indicated plasmid constructs using X-tremeGENE HP transfection reagents (Roche, Basel, Switzerland) according to the manufacturer’s instructions. For subsequent analysis following CRISPR-Cas9 mediated telomere removal, SH-SY5Y cells or HEK-293T cells were replated on appropriate culture plates 24 h after lentiCRISPR-gRNA (telomere) transfection.