SiRNAs or anti-sense nucleotides (ASO) specifically targeting circPGR were purchased from RiboBio (siRNA targeting sequence: CCAGGGCAGCACAACUACU dTdT; ASO targeting sequence: GCCAGGGCAGCACAACUAC dTdT). miR-301a-5p mimic and inhibitor were also purchased from RiboBio. SiRNA transfections were performed using Lipofectamine 2000 (Invitrogen) according to the manufacturer's protocol. Plasmid transfections in HEK293T cells were performed using Polyethyleneimine (PEI, Polysciences) according to the manufacturer's protocol. Plasmids transfection in MCF7 cells were performed using Lipoplus reagent (Sagecreation) according to the manufacturer's protocol.

For Lenti-virus packaging: HEK293T cells were seeded in culture plates coated with poly-D-lysine (0.1% (w/v), Sigma, P7280) and transfected with lenti-viral vectors together with packaging vectors, pMDL, VSVG and REV, at a ratio of 10:5:3:2 using Polyethyleneimine (PEI, Polysciences) for 48 h according to the manufacturer's protocol. Virus were collected, filtered and added to MCF7 cells in the presence of 10 μg/mL polybrene (Sigma, H9268), followed by centrifugation for 30 min at 1,500 g at 37 °C. Medium was replaced 24 h later.

注意:以上内容是从某篇研究文章中自动提取的,可能无法正确显示。



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