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文献和实验Protein Syntheses in Cell Free Systems
LEVEL III Materials Suspension culture of fibroblast cells (1 liter) 35 mM Tris-HCl, pH 7.4, 140 mM NaCl (TBS buffer) 10 mM Tris-HCl, pH 7.5, 10 mM KCl, and 1.5 mM magnesium acetate (TBS-M) 10X TBS-M
phase to a new 1.5 ml Eppendorf tube and add 50 µl 4 M ammonium acetate and 250 µl ice cold 100% ethanol. Invert the tube to mix and incubate for 30 minutes at -70℃. Spin in a microfuge for 15 minutes at 4℃. 6.Carefully remove the supernatant and add
RNase Protection Assay (Bowtell Lab)
0.5ul 200mM DTT 0.5ul appropriate RNA polymerase Incubate for 90-120min at 41�. 5. Add 0.5ul RNAsin and 0.5ul RNAse free DNAse (5mg/ml). Incubate for 15min at 37�. 6. Add 10ug tRNA and 100ul DDW. Phenol/CHCl3 extract and precipitate with 50ul 5M NH
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