Pre mir 19a

The cells were seeded on six-well plates and transfected the following day using Lipofectamine 2000 (Invitrogen, CA) based on the manufacturer's protocols. For each well, an equal dose (100 pmol) of pre-miR-control, pre-miR-19a, anti-miR-control, or anti-miR-19a (GenePharma, China) was added. The cell culture supernatants or the cells were harvested 48 h after transfection for further experiments.
A green fluorescence protein (GFP)-labeled miR-19 inhibitor lentivirus was purchased (Asia-Vector Biotech., China) and transfected into PAN02 cells according to the manufacturer's instructions for further homograft use.

MiRNA overexpression was achieved by transfecting cells with a miRNA mimic, whereas knockdown was achieved by transfecting cells with a miRNA inhibitor. Synthetic RNA molecules, including pre-miR-19a and pre-miR-19b (miRNA mimics), anti-miR-19a and anti-miR-19b (miRNA inhibitors), and scrambled negative control RNAs (pre-miR-control and anti-miR-control), were purchased from GenePharma (Shanghai, China). A549, H1975, and HCC827 cells were seeded in 6-well plates and transfected with Lipofectamine 2000 on the following day, when the cells were approximately 80% confluent. For miRNA overexpression, equal amounts of pre-miR-19a (100 pmol), pre-miR-19b (100 pmol) or pre-miR-19a/b (50 pmol each) were used. For miRNA knockdown, equal amounts of anti-miR-19a (100 pmol), anti-miR-19b (100 pmol) or anti-miR-19a/b (50 pmol each) were used. After 6 h, the medium was changed to DMEM or RPMI-1640 supplemented with 2% FBS. The cells were harvested 48 h after transfection for total RNA or protein isolation, respectively.