For functional knock-down of HDAC6, an HDAC6-targeting small interfering RNA (siRNA; siHDAC6, Sigma) and the universal negative control siRNA were transfected into hDPCs. Cells were seeded into 6-well plates and then transfected with siRNA using Lipo2000 (Invitrogen, Thermo Fisher Scientific, Waltham, MA, United States) according to the manufacturer's instructions (Tao et al., 2019 (link)). Knockdown efficacy was confirmed by western blot. Odontoblastic differentiation was induced 48 h after transfection. For the tracing of autophagic flux, mRFP-GFP-LC3 plasmid (plasmid 21074; Addgene) was transfected into hDPCs following transfection with siRNA prior to the induction of odontoblastic differentiation using Lipo2000 according to the manufacturer's instructions(Kimura et al., 2007 (link)).
Lipo2000
Manufactured by Addgene
Sourced in United States
Lipo2000 is a benchtop instrument used for the transfection of cells with nucleic acids, such as plasmid DNA or RNA. It employs a lipid-based delivery method to efficiently introduce genetic material into a variety of cell types.
Automatically generated - may contain errors
2 protocols using lipo2000
1
HDAC6 Knockdown in Human Dental Pulp Cells
2
Investigating Gene Expression Modulation
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TRIM37 siRNA (siR-1, 5'-GGAGAAGAUUCAGAAUGAATT-3'; siR-2, 5'-CCAGUAGUUUACUAGACAUTT-3'; siR-3, 5'-GCCUUGAUACAUGGCAGUATT-3'), METTL3 siRNA (siR#1, 5'-GCUGCACUUCAGACGAAUUTT-3'; siR#2, 5'-GGAUACCUGCAAGUAUGUUTT-3'), and IGF2BP2 siRNA (siR&1, 5'-CCCAGUUUGUUGGUGCCAUTT-3'; siR&2, 5'-GCGAAAGGAUGGUCAUCAUTT-3') were synthesized by Genepharm (Shanghai, China). For ectopic expression of TRIM37, GPX4, METTL3, or IGF2BP2, the coding sequences were cloned into pcDNA3.1 plasmids (Addgene, USA).
Transfections were performed with Lipo2000.
Transfections were performed with Lipo2000.
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