Method for sifting tumour chemotherapy medicament sensitive gene with RNA interference library
A technology of RNA interference and gene screening, applied in the field of biomedicine, can solve problems such as the limitation of tumor chemotherapy effects, the inability to conduct research, development and utilization of drug-sensitive genes, and the reduction of expression means
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[0050] 1. Using RNA interference library DNA to transfect Phoenix retrovirus packaging cells
[0051] Day 1: Preparing Phoenix Retroviral Packaging Cells
[0052] 1. 18-24 hours before transfection, press 3×10 6 / 10 cm plate inoculation of Phoenix cells onto the plate;
[0053] 2. Let the cells adhere to the wall for 10-24 hours;
[0054] 3. When the cell density reaches 2 / 3, a 10 cm plate has about 5×10 6 cell. At this time, the packaging cells are most easily transfected and produce the highest titer of virus at 37°C;
[0055]*Medium: DMEM, 10% fetal bovine serum, 1% penicillin-streptomycin, 1% glutamine.
[0056] Day 2: Transfection
[0057] 1. Prepare the DNA for transfection and dissolve it in HEPES solution;
[0058] 2. About 5 minutes before transfection, add 25 μM chloroquine to the plate (the storage concentration of chloroquine is 50 mmol);
[0059] * Chloroquine inhibits DNase in lysosomes by neutralizing the pH inside intracellular transport microvesicles; ...
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