Mouse L-type fibroblasts were used to perform the conditions eubiquitously, but can be applied to almost any cell with only minor modifications. It is critical to optimize the method according to the cell type under study.

Operation method

basic program

Materials and Instruments

DNA
TBS DMEM Dextran DMSO PBS
Incubator Tubes

Move

1. Inoculate about 5×10⁵ mouse L-type fibroblasts/10 cm dish and culture for 3 days to 30%~50% confluence.2. For each dish to be transfected, precipitate 4 μg of plasmid DNA to be transfected with ethanol, dry the precipitate in a tissue culture cabinet, and resuspend in 40 μl of TBS.

3. Aspirate off the culture solution and wash the petri dishes with 10 ml of 1×PBS, then wash once with 4 ml of complete DMEM-10NS.
4. Slowly (while shaking the tube continuously) add the DNA to 80 μl of pre-warmed 10 mg/ml DEAE-dextran/TBS. Add 120 μl of DNA/DEAE-dextran dropwise to each petri dish using a 200 μl pipette to evenly cover the entire surface of the dish, swirling gently to make it uniform until a consistent red color is obtained. Incubate in a humidified _CO2 incubator for 4 h (may be shorter for some cell types).
5. Aspirate the culture medium containing DNA/DEAE-dextran from the culture blood. Add 5 ml 10% DMSO/PBS to shock the cells. Leave at room temperature for 1 min, aspirate DMSO, wash once with 5 ml 1×PBS and aspirate 10 ml complete culture medium per dish.

6. Cultivate the cells and analyze at the appropriate time.

For more product details, please visit Aladdin Scientific website.

https://www.aladdinsci.com/